FIELD OF THE INVENTION

The present disclosure provides a digestate active filler composition suitable for use in asphalt compositions and asphalt compositions comprising the digestate active filler composition. The digestate active filler composition is obtainable from waste treatment processes wherein waste is subjected to enzymatic and/or microbial treatment followed by anaerobic digestion of the liquid fraction thus obtained.

BACKGROUND OF THE INVENTION

Active fillers such as mineral filler, cement and lime in asphalt mixture may be important components of an asphalt composition as it plays an important role in stiffening and toughening an asphalt binder. In addition to affecting the mechanical properties of asphalts, mineral fillers are also important with respect to stripping or moisture damage. In asphalt bitumen layers active fillers are often added to improve the quality of the asphalt and provide e.g., an arming effect, since the softening point is increased, and the risk of cracking thereby decreased. The filler can stabilize the bitumen, since it binds part of the bitumen oils, which could otherwise evaporate and make the bitumen harder. Furthermore, the filler can decrease the effect from UV-light, thus the bitumen decomposes at a lower rate. The addition of filler may also decrease the flammability of bitumen. It is well known that the physical properties of the mixtures and binding components in an asphalt composition is highly dependent on type and concentration of active filler (Diab and Enib, 2018). Normally, the ration between filler and binder such as bitumen is from 0.6 - 1 .2.

Most road pavements consist of several layers of different materials which in combination make the road strong and durable. Asphalt pavements are frequently described as flexible pavements, implying their ability to absorb the stresses imposed by traffic and weather without cracking. An example of the component layers of such pavements is illustrated in Figure 1. The subgrade is typically the natural soil, on old roads usually well compacted by traffic, on new roads carefully shaped and compacted to the appropriate level and profile. Subgrade improvement may be possible by treatment of soils with lime, cement and Ground Granulated Blastfurnace Slag (GGBS) or by adding a ‘capping layer’ of lower quality aggregate. The sub-base is the lowest layer, put down to help build up the strength of the pavement. It also provides a working platform for the machinery used in laying and compacting the layers above. It is usually made from crushed stone and/or gravel.

The base is typically the main component of an asphalt pavement and provides most of the strength and load distributing properties of the pavement. For very lightly trafficked roads, car parks and pedestrian footways, it is usually made from graded crushed stone and/or hardcore and/or it may be crushed stone bound with a small proportion of cement (cement-bound granular base or lean-mix concrete) and other Hydraulically Bound Materials (HBMs). For most roads and areas carrying heavy vehicles, however, an asphalt base is used to provide a pavement of high strength and durability, to achieve the desired load-bearing capacity and absorb traffic loads so that the underlying subgrade is not deformed. The binder course typically further contributes to the strength of the pavement, and at the same time provides an even, well-regulated surface to carry the uppermost layer of the pavement. The surface course typically provides an even and weather-resistant surface which can withstand the abrasive forces of traffic and provide appropriate skid resistance for the particular circumstances. Roads are exposed to particularly high stresses, e.g., when the water contained in the pavement structure begins to freeze. Water expands when freezing, which can lead to frost damage that will sooner or later have an impact also on the road surface. This is prevented by a so- called frost blanket which usually consists of a mixture of gravel and sand, supplemented by crushed mineral aggregate. When compacted, these layers of frost-resistant materials conduct water away from the upper pavement layers, reducing tensions very effectively at the same time.

Environmentally friendly waste processing methods using biologically based technology, wherein waste comprising organic matter is converted to energy have been developed, however, depending on the composition and source of the waste, some of the waste fractions cannot be converted by enzymes and/or microorganism into energy or other valuable products and needs to be disposed. One such fraction is digestate, which is a solid remaining after anaerobic digestion process, of various substrate including different biomass and e.g., bioliquid obtained from enzymatic and/or microbial degradation of waste comprising organic matter.

Anaerobic digestion (AD) is a series of biological processes in which microorganisms break down biodegradable material in the absence of oxygen. One of the end products is biogas, which can be combusted to generate electricity and heat, or can be processed into renewable natural gas and transportation fuels. A range of anaerobic digestion technologies exists in the state of the art for converting waste, such as municipal solid waste (MSW), municipal wastewater solids, food waste, high strength industrial wastewater and residuals, fats, oils and grease, and various other organic waste streams into biogas. Apart from the production of valuable output, AD also produces a digestate as for example provided in EP 3 569657 A1 , comprising the waste product from anaerobic digestion.

Whereas there is a need to dispose waste products or re-use them in an environmentally beneficial way, such as adding waste products to asphalt compositions, this is only feasible if the quality of the asphalt composition will not be compromised. An adequate selection of the materials and asphalt mixtures used in the construction of road structures exerts a fundamental role in the performance, durability, and functionality of the transport infrastructure. An optimal asphalt mixture design would guarantee sufficient bearing capacity, enough flexibility to counteract the initiation of cracking at low stress levels, a good interaction between the components, and the capacity of resist fatigue associated with both traffic loads and thermal stresses. Fatigue phenomena could give way to the progressive cracking and breakage of the asphalt mixture layers of the pavement structure, which is one of the main reasons behind structural failure in pavements. Thus, using waste or by-products as filler in asphalt compositions may compromise any of these required features of the asphalt composition, and particularly, if the filler is an active filler, meaning that the filler influences the performance of the asphalt. Of course, if the asphalt composition is improved on one or more ways when the active filler is added, this would be advantageous.

Anaerobic digestate that can be obtained from anaerobic digestion of organic matter such as liquified household waste has been added to asphalt compositions previously (EP 3 569 657 A1) to provide asphalt mixture compositions wherein anaerobic digestate additive was partly replacing a polymer modified binder. It was found in EP 3 569 657 A1 that partly replacement of the binder increased the softening point of the binder in the asphalt composition.

The present invention provides a specific digestate active filler composition and use thereof in asphalt compositions. Adding the digestate active filler composition to asphalt compositions was surprisingly found to lower the softness, improve anti-ageing and rejuvenation in asphalt compositions while upholding sufficient tensile strength.

When used as active filler the digestate has a particle size below 2 mm and at least 30% of the particles of in the active filler composition is 0.063 mm or less. Some of the components comprised by the digestate active filler are known to have potential beneficial impact on bitumen in asphalt (e.g. anhydrous carbon fibres, lignin, plastic polymers, fatty acids, inorganic components comprised in the ash fraction) whereas other components comprised by the digestate active filler are expected to be inert (e.g. proteins). However, the potential positive impact of the individual components on the properties of the asphalt composition depends on the amounts added to the bitumen or asphalt composition. It was surprisingly found here that digestate with its specific components and the combined amounts of these components could be added as active filler to asphalt compositions. Moreover, it was very surprising that in 2 - 3 times the amount of the digestate active filler could be added to an asphalt composition compared to the amount of digestate that had previously been added to an asphalt composition. The positive impact of adding the digestate on improving the asphalt properties was found when replacing filler material of the asphalt compositions, with the digestate active filler - hence the term active filler. It was found that replacing part of natural filler such as limestone with digestate active filler did not have a negative impact on the properties of the asphalt composition. The fact that a higher amount of digestate active filler can be added to asphalt compositions without degrading the properties of the asphalt composition has a beneficial impact on the CO2 footprint by reducing the amount of filler that would otherwise be required.

When replacing natural and similar filler materials with digestate active filler in asphalt compositions the CO2 footprint of the asphalt composition is significantly improved in particular when the digestate is obtained from waste. Digestate active filler contains biogenic carbon and by use of recognized Life Cycle Assessment methods the digestate active filler provides a net negative CO2 emission making it an environmentally sustainable alternative to replace conventional fillers, such as limestone, in the amounts now found suitable.

SUMMARY OF THE INVENTION

The present invention relates to a digestate active filler composition and to the use thereof in an asphalt composition and to asphalt compositions comprising the active filler composition.

In a first aspect, the invention provides a digestate active filler dry matter composition comprising:

Active filler composition comprising:

Anhydro carbohydrates 10 - 14 wt%

Acid resistant compounds 15 - 24 wt% wherein the acid resistant compounds are determined according to NREL/TP-510-42618 and comprise 30-70% protein and protein complexes estimated from nitrogen and ammonia content, 15- 35 % lignin and lignin derivatives; and 0.5-45% plastic derived compounds

Ash, measured by thermogravimetric analysis at 550°C 40 - 62 wt% wherein the ash comprises 10-75% calcium and/or calcium salts

Fatty acids 0.1 - 5 wt%

Other 0 - 10 wt% wherein the total sum of the components is 100% by weight of the active filler composition, and wherein at least 99% of the particles in the active filler composition have a particle size of 2 mm or less, and wherein at least 30% of the particle size of the particles in the active filler composition is 0.063 mm or less.

This filler composition is obtainable from many different sources.

In a second aspect of the invention, the active filler composition is obtained from a method comprising: a. Subjecting a waste fraction to anaerobic digestion b. Separating the solid fraction from the liquid fraction of the digestate obtained in step a) c. Optionally subjecting the solid digestate fraction obtained in step b) to anti-lumping treatment and/or passing digestate through 2 mm sieve d. Optionally subjecting the digestate obtained in c) to drying and/or milling to reduce particle size

The method may further comprise two initial steps: i. Subjecting waste to enzymatic and/or microbial treatment ii. Subjecting the treated waste from step i) to one or more separation step(s) whereby a bioliquid fraction and a solid fraction is provided, wherein the bioliquid fraction obtained from step ii) is the waste fraction subjected to anaerobic digestion according to claim 6 step a).

The waste may be one or more of sludge, seaweed or the waste may be one or more of sorted MSW, unsorted MSW, agricultural waste, industrial waste, waste fractions from paper industry; waste fractions from recycling facilities; waste fractions from food or feed industry; waste fraction from the medicinal or pharmaceutical industry; waste fractions from hospitals and clinics, waste fractions derived from agriculture or farming related sectors; waste fractions from processing of sugar or starch rich products; contaminated or in other ways spoiled agriculture products, and garden refuse or any other waste containing biodegradable/biogenic material that can be digested in anaerobic digestion plant.

The active filler composition is in one aspect of the invention an anaerobic digestate that is obtainable as a solid fraction from an anaerobic digestate process being the fraction of the anaerobic digestate that consist of particles larger than 2 mm and particles smaller than 2 mm (i.e. , 2 mm and less). In one preferred aspect of the invention the active filler composition is an anaerobic digestate consisting of particles of approximately 2 mm and less, where the anaerobic digestate is obtained from an anaerobic digestion process, where the substrate of the anaerobic digestion process is the bioliquid obtained from enzymatically and/or microbial degradation of waste such as MSW.

The size of the particles in the composition can be adjusted for example by subjecting the composition to size separation and/or milling for reducing the particle size. The means for reducing the particle size can be any means available in the art.

The composition of the invention may be used in the binder course, regulating course and/or in the surface course of asphalt composition.

Thus, in a third aspect the present invention relates to the use of the digestate active filler composition in the binder course, regulating course and/or in the surface course of asphalt compositions.

It is shown herein that the digestate active filler composition provides one or more of a softening effect, and anti-ageing effect and a rejuvenation effect when applied in the regulating course, binder course or surface course of an asphalt composition. Thus, in a fourth aspect, the present invention relates to the use of the digestate active filler composition for one or more of softening, anti-ageing and rejuvenation of the binder course, regulating course and/or the surface course of an asphalt composition.

In a fifth aspect, the present invention provides asphalt binder course compositions comprising up to 100% of digestate active filler composition according to the invention of the total amount by weight of fine aggregate components in the binder course composition.

In a sixth aspect, the present invention provides asphalt surface course composition comprising up to 100% of digestate active filler composition according to the invention of the total amount by weight of filler aggregate components in the surface course composition.

In a seventh aspect, the present invention provides asphalt compositions comprising up to 100% of digestate active filler composition according to the invention of the total amount by weight of fine aggregate or filler aggregate components in the surface course, regulating course and/or in the binder course composition.

BRIEF DESCRIPTION OF THE FIGURES

Figure 1 : A - Graph showing particle size distribution in milled samples of active filler compositions, B - Graph showing particle size distribution in un-milled samples of active filler compositions Figure 2: Schematic drawing of devise for measuring tensile strength

Figure 3: Schematic drawing of devise for ball and ring measuring

Figure 4: Diagrams showing the penetration (a), softening point (b), and penetration index (specification limit: -1.5) (c) of reference asphalt samples compared with samples comprising active filler composition for bitumen extracted from fresh and aged samples

Figure 5: Diagrams showing the indirect tensile strength for A: dry and wet samples, B: ratio, C: as a function of sampling date, D: ratio as a function of sampling date.

Figure 6: Diagrams showing A: cycles to failure, B: cumulative axial strain.

Figure 7: A - Master curves showing shear stress modulus (G, mPa) as a function of angular frequency (rad/s) in fresh and aged samples of extracted bitumen, B - Master curves showing phase shift angle (p, °) as a function of angular frequency (rad/s) in fresh and aged samples of extracted bitumen, C - Master curves with confidence intervals showing shear stress modulus (G, mPa) and phase shift angle (p, °) as a function of angular frequency (rad/s) in fresh and aged samples of extracted bitumen.

Figure 8: Box and whiskers plot for complex shear modulus ratio between samples of unaged and aged bitumen.

Figure 9: Diagram of organic carbon (by DUMAS, %) and estimated protein content (wt% of dry matter) of digestate samples on different dates.

Figure 10: HSEQ NMR spectra of aromatic region and of aliphatic region of alkaline lignin extracts from dried digestate samples.

Figure 11 : Photos of A: 1 mm sieve fraction after wet sieving, B: 1 mm sieve fraction of after dry sieving, C: 2 mm sieve fraction after wet sieving, D: 2 mm sieve fraction after dry sieving.

Figure 12: Photos of a): Unsorted material from dried 2 mm sieve fraction, b) Sorted and identified material from dried 2 mm sieve fraction.

Figure 13: HSQC NMR spectra of digestate DCM extracts showing carbon stretch (SC, PPM) on the F1 axis and hydrogen-stretch (5H, PPM) on F2 axis.

Figure 14: Diagrams showing response (pA) over time (minutes) of GC-FID analysis of FAME derivatized DCM extracts at A: ambient temperature extract, and B: reflux extract. Figure 15: Diagrams showing relative abundance over time (minutes) from GC-MS analysis of derivatized DCM extract of A: ambient temperature extract, B: reflux extract, and C:underivatized combined DCM extracts.

DEFINITIONS

Acid resistant compounds is the fraction measured as Klason-lignin after analysis according to Sluiter et al 2008, revised 2012 (NREL/TP-510-42618): Determination of Structural Carbohydrates and Lignin in Biomass when performing the analysis on a material that is not solely a lignocellulosic material. Samples with a LO c higher than 10 wt% of dry matter can be biased due to interference with appropriate acid concentration or may catalyze side reactions. Interferece with acid concentration or side reactions will lead to over estimation of acid resistant compounds. Remaining extractives such as waxes and oils can also cause positive intereferense on the analysis according to A. Sluiter et al., Methods for Biomass Compositional Analysis, Max Planck Library for the History and Development of Knowledge, Proceedings 2, 2013. Further, samples containing protein will bias acid insoluble compounds which can be countered by performing an independent nitrogen analysis and estimating the protein content and subtracting the protein estimate from the acid resistant compounds.

The acid resistant compounds comprise approx. 15-24 wt% such as 18-24 wt% of the dry matter of the sample, where the protein content is estimated to be approximately 30-70 %, plastic compounds is estimated to be 0.5-45%. Lignin is found by py-GC-MS to be 15-35 %, which is probably underestimated due to only analysing some constituents in lignin and pyrolysis also being prone to experience negative interferences when used on samples with a high ash content.

A digestate active filler is in the present context a digestate as defined below having active filler properties. A digestate active filler is an active component, improving the properties of an asphalt composition comprising it. A digestate active filler has the ability to increase the resistance of particles to move within the mix matrix and/or works as an active material when it interacts with the asphalt composition to change the properties of the mastic. The term digestate active filler comprises in the context of the present invention “fine aggregates” and “filler aggregates” as defined below.

Aggregate is granular material used in construction. Aggregate may be natural, manufactured or recycled. Natural aggregate is from mineral sources which has been subjected to nothing more than mechanical processing. Manufactured aggregate is of mineral origin and results from an industrial process involving thermal or other modification. Recycled aggregate results from processing of inorganic material previously used in construction. Aggregate size is designated in terms of lower (d) and upper (D) sieve sizes expressed as d/D. This designation accepts the presence of some particles which are retained on the upper sieve (oversize) and some which pass the lower sieve (undersize). The lower sieve side (d) can be zero. Larger aggregate sizes with D less than or equal to 45 mm and d greater than or equal to 2 mm is designated coarse aggregate. Aggregate sizes with D less than or equal to 2 mm and containing particles which mostly are retained on a 0.063 mm sieve are designated fine aggregate. Filler aggregate is the aggregate, most of which passes a 0.063 mm sieve, which can be added to construction materials to provide certain properties.

Ash content is the inorganic residue after dry oxidation at a defined temperature, typically 550°C, 575°C or 900°C. In the context of the present invention the ash content is the inorganic residue after dry oxidation at 550 or 575°

For the purpose of the present invention, asphalt composition, asphalt concrete, asphalt mixture composition, asphalt mixture, asphalt mix, asphalt pavement or simply asphalt means a composition comprising at least one aggregate, at least one filler, optionally at least one additive and at least one binder, such as bitumen and/or other, in varying amounts and relative percentages. The term ‘asphalt’ is herein also used to describe the wide range of mixtures of, at least, bituminous binder and aggregate individually known as Warm mix asphalt (WMA), Hot Rolled Asphalt (HRA), Stone Mastic Asphalt (SMA), Thin Surfacings, Mastic Asphalt, Asphalt Concrete etc., which are available for use in constructing and maintaining paved areas. Asphalt pavements are frequently described as flexible pavements, implying their ability to absorb the stresses imposed by traffic and weather, without cracking. An asphalt mixture composition according to the present invention is suitable for building roads, pavements and paved areas, roofing, vehicle parking areas, housedrives, footways, recreation areas such as tennis courts or playgrounds, agricultural uses such as farm roads or animal cubicles, airfields, runways and access roads, hard standings, storage areas, hydraulic applications such as dam construction, coastal protection or other.

Anaerobic digestion refers to the biological processes in which microorganisms such as archaea break down biodegradable material in the absence of oxygen. One of the end products may be biogas, which can e.g., be combusted to generate electricity and/or heat. Biogas can also be used, either directly or after upgrading, as renewable natural gas and/or transportation fuels. Biogas can be injected into a natural gas and/or biogas grid.

Anhydro carbohydrates are intramolecular ethers formed by the loss of the elements of water from neighbouring hydroxyl groups of a sugar. Anhydro-sugars are bicyclic monosaccharides having 1 ,6- and 1 ,5-anhydro rings for hexoses and 1 ,4- and 1 ,5-rings for pentoses, respectively, which may be derived from the corresponding monosaccharides or polysaccharides such as cellulose, xylose, arabinose, amylose, and mannan. The anhydro carbohydrates found in degradation products from waste are often degradation products from cellulose, hemicellulose, pectin, food products and comprises for example arabinose, galactose, glucose, mannose, arabinose, glucoronic acid, xylose, galacturonic acid, uronic acid and rhamnose. In the present invention, structural carbohydrates are determined according to Sluiter et al 2008, revised 2012, (NREL/TP-510-42618): Determination of Structural Carbohydrates and Lignin in Biomass as anhydrous xylose, arabinose or galactose.

Binder when used in asphalt mixture compositions binds the aggregate particles into a cohesive mixture, whilst also lubricating the particles when hot to assist in compaction. According to the type of mixture and its end use, the amount of binder used will typically vary between 3 and 9 percent by mass of the mixture. Bituminous binders are characterised by being the residue from distilling raw oil and are black, sticky and thermoplastic, i.e., they become softer and more fluid when they are heated and harden again when they cool. Bituminous binders are named after their penetration at 25°C. A Bitumen 40/60 is a bitumen with a penetration of 4-6 mm penetration at 25°C and a bitumen 70/100 is a bitumen with at penetration of 7-10 mm at 25°C measured according to EN 1426:2015.

Biodegradable matter refers to organic matter that can be partly or completely degraded into simple chemical compounds such as mono-, di- and/or oligo-saccharides, amino acids and/or fatty acids by microorganisms and/or by enzymes. Biodegradable matter is generally organic material that provides a nutrient for microorganisms, such as mono-, poly- or oligosaccharides, fat and/or protein. These are so numerous and diverse that a huge range of compounds can be biodegraded, including hydrocarbons (oils), polycyclic aromatic hydrocarbons (PAHs), polychlorinated biphenyls (PCBs) and pharmaceutical substances. Microorganisms secrete biosurfactant, an extracellular surfactant, to enhance this process.

Bioliquid is the liquefied and/or saccharified degradable components obtained by enzymatic treatment of waste comprising organic matter. Bioliquid also refers to the liquid fraction obtained by enzymatic treatment of waste comprising organic matter once separated from non-fermentable solids. Bioliquid comprises water and organic substrates such as protein, fat, galactose, mannose, glucose, xylose, arabinose, lactate, acetate, ethanol and/or other components, depending on the composition of the waste (the components such as protein and fat can be in a soluble and/or insoluble form). Bioliquid comprises also fibres, ashes and inert impurities. The resulting bioliquid comprising a high percentage of solubles provides a substrate for gas production, a substrate suitable for anaerobic digestion e.g., for the production of biogas.

“Digestate”, sometimes called, and used interchangeably with “anaerobic digestate”, “AD digestate” or “solid digestate” is the residual output from an anaerobic digestion (AD). Usually, the digestate has alkaline pH and comprises mainly water, but also suspended solids and dissolved matter such as salts which may include both inorganic salts and organic salts. It is the material remaining after anaerobic digestion of a biodegradable feedstock such as bioliquid from an enzymatic and/or microbial treatment of waste or other substrates suitable for anaerobe digestion. The digestate may advantageously be dewatered by separation means, such as filters, sedimentation tanks or the like into "dewatered digestate" or “solid digestate” and "reject water".

Dry matter, also appearing as “DM”, refers to total solids, both soluble and insoluble, and effectively means "non-water content." Dry matter content is measured by drying at approximately between 60 to 105°C until constant weight is achieved. In a preferred embodiment, dry matter content is measured by drying at approximately 105 °C. The lower temperature range is used when the analysis substrate contains volatile compounds which may escape when boiling water and decrease the analysis result accuracy.

Filler or filler aggregate refers to a fraction of a mineral most of which passes a 63 pm sieve. Main function of a filler is that of filling voids in coarse aggregates, which intensifications the density, stability and toughness of a conventional bituminous paving mix. Another is the formation of a fillerasphalt mastic in which the particles of dust either may be individually coated with asphalt or are fused into the bitumen in mechanically and colloidal suspension. Excess amount of fillers leads to, brittleness and tendency to cracking. Deficiency of filler leads to increase void content, lower stability and softens the mix.

Hydrolysis is meant to be related to the context wherein the waste, such as municipal solid waste material is hydrolysed to break down cellulose and/or hemicellulose and other substrates to fermentable sugars, such as glucose, cellobiose, xylose, xylulose, arabinose, mannose, galactose, and/or soluble oligosaccharides (also known as saccharification). The hydrolysis is performed enzymatically by one or more enzyme compositions in one or more stages. In the hydrolysis step, the waste, such as municipal solid waste material, e.g., pre-treated, is hydrolysed to break down proteins and lipids (e.g., triglycerides) found in the waste. The hydrolysis can be carried out as a batch process or series of batch processes. The hydrolysis can be carried out as a fed batch or continuous process, or series of fed batch or continuous processes, where the waste, such as municipal solid waste material is fed gradually to, for example, a hydrolysis solution comprising an enzyme composition. The hydrolysis may be continuous hydrolysis in which a waste material, such as municipal solid waste (MSW), and an enzyme composition are added at different intervals throughout the hydrolysis and the hydrolysate is removed at different intervals throughout the hydrolysis. The removal of the hydrolysate may occur prior to, simultaneously with, or after the addition of the cellulosic material and the cellulolytic enzymes composition.

Municipal solid waste (MSW) refers to waste fractions which are typically available in a city, but that need not come from any municipality per se, i.e. , MSW refers to every solid waste from any municipality but not necessarily being the typical household waste - could be waste from airports, universities, campus, canteens, general food waste, among others. MSW may be any combination of one or more of cellulosic, plant, animal, plastic, metal, or glass waste including, but not limited to, any one or more of the following: Garbage collected in normal municipal collections systems, optionally processed in a central sorting, shredding or pulping device, such as e.g., a Dewaster® or a reCulture®; solid waste sorted from households, including both organic fractions and paper rich fractions; Generally, municipal solid waste in the Western part of the world normally comprise one or more of: animal food waste, vegetable food waste, newsprints, magazines, advertisements, books and phonebooks, office paper, other clean paper, paper and carton containers, other cardboard, milk cartons and alike, juice cartons and other carton with alu-foil, kitchen tissues, other dirty paper, other dirty cardboard, soft plastic, plastic bottles, other hard plastic, non-recyclable plastic, yard waste, flowers etc., animals and excrements, diapers and tampons, cottonsticks etc., other cotton etc., wood, textiles, shoes, leather, rubber etc., office articles, empty chemical bottles, plastic products, cigarette buts, other combustibles, vacuum cleaner bags, clear glass, green glass, brown glass, other glass, aluminium containers, alu-trays, alu-foil (including tealight candle foil), metal containers (-AI), metal foil (-AI), other sorts of metal, soil, rocks, stones and gravel, ceramics, cat litter, batteries (botton cells, alkali, thermometers etc.), other non-combustibles and fines.

Model MSW can be prepared to mimic the composition of real municipal solid waste. A typical model MSW consisting of 3 main fractions:

10-60% vegetable fraction prepared from a variety of vegetables and fruits

5-30% protein/fat fraction (animal origin) prepared from a variety of non-processed, processed and/or cooked meats and

30-80% cellulosic fraction prepared from a variety of paper/cardboard and cotton sources.

Protein content in a digestate from a biogas process is estimated according to J. Liebetrau and D. Pfeiffer, Biomass energy use, Vol. 7, collections of Methods for Biogas in biogas digestates with a solid content up to 7 wt%, from the nitrogen content subtracted the ammonia and nitrate and nitrite content from the liquid portion of the sample determined by the DUMAS method multiplied by a factor of 6.25.

Rut depth is the measure of the depth of the rut a wheel makes in the asphalt after running back and forth a set number of times and is thus the reduction in the thickness of a test specimen asphalt mixture composition, in millimetres or percentage, relative to the specimen original thickness, caused by repeated passes of a loaded wheel according to EN 12697-22:2003+A1 :2007. Softening point of a binder is the temperature at which a material under standardised test conditions described in EN 1427:2015 attains a specific consistency.

Solid/liquid separation refers to an active mechanical process, and/or unit operation(s), whereby liquid is separated from solid by application of some force through e.g., pressing, centrifugation, sedimentation, decanting or the like. Commonly, a solid/liquid (s/l) separation provides a liquid and solid fraction.

Sorted waste (and "sorted MSW') as used herein refers to waste, such as MSW, in which approximately less than 30%, preferably less than 20% and most preferably less than 15% by weight of the dry weight is not biodegradable material.

Stiffness modulus is a parameter expressing the relationship between stress and strain when submitting a linear viscoelastic material to a sinusoidal load wave. Stiffness modulus can be determined according to EN 12697-26:2012.

Unsorted waste (and “unsorted” MSW”) refers to waste that is not substantially sorted into separate fractions such that organic material is not substantially separated from plastic and/or other inorganic material, notwithstanding removal of some large objects or metal objects and not withstanding some separation of plastic and/or other inorganic material may have taken place. The terms "unsorted waste" (or "unsorted MSW'), as used herein, refers to waste comprising a mixture of biodegradable and non-biodegradable material in which 15% by weight or greater of the dry weight is non- biodegradable material. Waste that has been briefly sorted yet still produce a waste (or MSW) fraction that is unsorted. Typically, unsorted MSW may comprise organic waste, including one or more of food and kitchen waste; paper- and/or cardboard-comprising materials; recyclable materials, including glass, bottles, cans, metals, and certain plastics; burnable materials; and inert materials, including ceramics, rocks, and debris. The recyclable material might be before or after source sorting.

Waste comprises, sorted and unsorted MSW, agriculture waste, hospital waste, industrial waste, e.g., waste fractions derived from industry such as restaurant industry, food processing industry, general industry; waste fractions from paper industry; waste fractions from recycling facilities; waste fractions from food or feed industry; waste fraction from the medicinal or pharmaceutical industry; waste fractions from hospitals and clinics, waste fractions derived from agriculture or farming related sectors; waste fractions from processing of sugar or starch rich products; contaminated or in other ways spoiled agriculture products such as grain, potatoes and beets not exploitable for food or feed purposes; or garden refuse. DETAILED DESCRIPTION OF THE INVENTION

The present invention provides in a first aspect an active filler composition comprising:

Anhydro carbohydrates 10 - 14 wt%

Acid resistant compounds 15 - 24 wt%, wherein of the acid resistant compounds are determined according to NREL/TP-510-42618 and comprise 30-70% protein and protein complexes, 15-35% lignin and lignin derivatives; and 0.5-45% plastic derived compounds

Ash, measured by thermogravimetric analysis at 575°C 40 - 62 wt%, wherein the ash comprises 10-75% calcium and/or calcium salts

Fatty acids or other apolar components 0.1 - 5 wt%

Other 0 - 10 wt%, wherein the total sum of the components is 100% by weight of the active filler composition, and wherein at least 99% of the particles in the active filler composition have a particle size of 2 mm or less, and wherein at least 30% of the particle size of the particles in the active filler composition is 0.063 mm or less.

In one embodiment, the active filler composition comprises:

Anhydro carbohydrates 10 - 14 wt%

Acid resistant compounds 18 - 24 wt%, wherein of the acid resistant compounds are determined according to NREL/TP-510-42618 and comprise 30-70% protein and protein complexes, 15-35% lignin and lignin derivatives, and 0.5-45% plastic derived compounds

Ash, measured by thermogravimetric analysis at 575°C 40 - 62 wt%, wherein the ash comprises 10-75% calcium and/or calcium salts

Fatty acids or other apolar components 0.1 - 5 wt%

Other 0 - 10 wt%, wherein the total sum of the components is 100% by weight of the active filler composition, and wherein at least 99% of the particles in the active filler composition have a particle size of 2 mm or less, and wherein at least 30% of the particle size of the particles in the active filler composition is 0.063 mm or less. In one preferred embodiment the active filler composition is a digestate, which is the residual output from an anaerobic digestion (AD). The substrate for the AD process may be any suitable material, comprising biodegradable components, such as sewage sludge, seaweed, manure, food waste etc. The substrate of the AD process may also be pre-treated waste, such as MSW. In one preferred embodiment the substrate for the AD-process is the bioliquid resulting from treatment of waste, such as MSW with enzymes and/or microorganisms, followed by separation of bioliquid from the nondegradable parts, such as plastic and metals. Thus, one embodiment of the invention relates to

A digestate active filler composition comprising:

Anhydro carbohydrates 10 - 14 wt%

Acid resistant compounds 15 - 24 wt%, wherein of the acid resistant compounds are determined according to NREL/TP-510-42618 and comprises 30-60% protein and protein complexes, 15-35 % lignin and lignin derivatives; and 2-75% plastic derived compounds

Ash, measured by thermogravimetric analysis at 550°C 40 - 67 wt% wherein the ash comprises 5-75% calcium and/or calcium salts

Fatty acids 0.1 - 5 wt%

Other 0 - 10 wt% wherein the total sum of the components is 100% by weight of the active filler composition, and wherein at least 99% of the particles in the active filler composition have a particle size of 2 mm or less, and wherein at least 30% of the particle size of the particles in the active filler composition is 0.063 mm or less.

In one embodiment, the digestate active filler composition comprises:

Anhydro carbohydrates 10 - 14 wt%

Acid resistant compounds 18 - 24 wt%, wherein of the acid resistant compounds are determined according to NREL/TP-510-42618 and comprise 30-70% protein and protein complexes, 15-35% lignin and lignin derivatives; and 0.5-45% plastic derived compounds

Ash, measured by thermogravimetric analysis at 575°C 40 - 62 wt%, wherein the ash comprises 10-75% calcium and/or calcium salts Fatty acids or other apolar components 0.1 - 5 wt%

Other 0 - 10 wt%, wherein the total sum of the components is 100% by weight of the active filler composition, and wherein at least 99% of the particles in the active filler composition have a particle size of 2 mm or less, and wherein at least 30% of the particle size of the particles in the active filler composition is 0.063 mm or less.

The term digestate active filler, means that the active filler is a digestate. As mentioned above digestate is a residual output from an AD process. An AD process may produce digestate from any substrate comprising biodegradable matter suitable for an AD process, such as but not limited to sewage sludge, food waste, seaweed.

Accordingly, the digestate can be obtained from a method comprising: a. Subjecting a substrate comprising biodegradable matter to anaerobic digestion b. Separating the solid fraction from the liquid fraction of the digestate obtained in step a) c. Optionally subjecting the solid digestate fraction obtained in step b) to antilumping treatment and/or passing digestate through 2 mm sieve d. Optionally subjecting the digestate obtained in c) to drying and/or milling to reduce particle size

Some type of substrate may need pre-treatment before subjected to the AD process and one embodiment relates to a digestate obtained from a method, as described above i.e. an AD process on substrate comprising biodegradable matter, further comprising two initial steps: i. Subjecting substrate comprising biodegradable matter to enzymatic and/or microbial treatment ii. Subjecting the treated substrate from step i) to one or more separation step(s) whereby a bioliquid fraction and a solid fraction is provided. wherein the bioliquid fraction obtained from step ii) is the fraction subjected to anaerobic digestion according to step a) above. In one preferred embodiment the digestate is derived from waste, such as MSW. Thus, in one embodiment the active filler is a digestate, wherein the digestate is obtained from a method comprising: a. Subjecting a waste fraction to anaerobic digestion b. Separating the solid fraction from the liquid fraction of the digestate obtained in step a) c. Optionally subjecting the solid digestate fraction obtained in step b) to antilumping treatment and/or passing digestate through 2 mm sieve d. Optionally subjecting the digestate obtained in c) to drying and/or milling to reduce particle size

Some type of waste may need pre-treatment before subjected to the AD process and one embodiment relates to a digestate obtained from a method, as described above i.e. an AD process on waste fraction, further comprising two initial steps: i. Subjecting waste to enzymatic and/or microbial treatment ii. Subjecting the treated waste from step i) to one or more separation step(s) whereby a bioliquid fraction and a solid fraction is provided. wherein the bioliquid fraction obtained from step ii) is the waste fraction subjected to anaerobic digestion according to step a) above.

Digestate is normally disposed and therefore reusing it for practical purposes is very beneficial. Particularly, reusing digestate as active filler in asphalt, this reduces the amount of natural filler that would otherwise be required in the asphalt composition reducing the CO2 required to provide such fillers. Digestate contains biogenic carbon and therefor when bound in asphalt it provides temporary and/or permanent carbon storage.

A digestate active filler composition comprises several degradation products from degradation of organic and inorganic matter, such as products obtainable from degradation processes of waste comprising organic matter. The anhydro carbohydrate fraction comprises intramolecular ethers formed by the loss of the elements of water from neighbouring hydroxyl groups of sugars. Anhydrosugars are bicyclic monosaccharides having 1 ,6- and 1 ,5-anhydro rings for hexoses and 1 ,4- and 1 ,5-rings for pentoses, respectively, and may be derived from the corresponding monosaccharides or polysaccharides such as cellulose, amylose, and mannan. The anhydro carbohydrates found in degradation products from waste are often degradation products from cellulose, hemicellulose, pectin, food products and waste obtained from household, agriculture or industrial processing of organic matter and comprises for example arabinose, galactose, glucose, mannose, arabinose, xylose, rhamnose, and uronic acids. The digestate active filler composition comprises between 10 and 14%, such as 10%, 11 %, 12%, or 13% anhydro carbohydrates by weight of the active filler composition. The anhydrous carbon fraction act as a stabilizer agent in asphalt mixtures According to C. Rodrigues and R. Hanumanthgari, The Shell Bitumen Handbook, 6 ^th ed., Chapter 8, p. 151 and G. Airey and A. Colop, ICE Manual of Construction Materials, Chapter 3, 2008, p 304. Addition of fibres such as cellulose or lignin are used e.g., to improve strength and decrease drainage of the binder from the asphalt mixture under transportation and placement due to viscosity increase. Inhibiting binder drainage is especially important for binder rich, soft and porous asphalts such as SMA mixtures.

The active filler composition moreover comprises a fraction of acid resistant compounds. This fraction comprises compounds that are difficult to degrade in natural systems and moreover resistant to degradation by acid treatment. Such compounds are for instance lignin and lignin-derived products or other phenol-based polymers or other acid resistant components. According to Landa and Gosselink, Stichting Wageningen Research, 2018, WO 2019/092278, the lignin and lignin-derived products act as chemical modifiers upon binders in asphalt mixtures. In plants, lignin acts a stabilizer and strengthens the structure, as cellulose fibres. Lignin has a structural similarity to the aromatic and asphaltene fraction of the binder, bitumen, in asphalt mixtures and has been found to have the ability to substitute some of the bitumen as well as provide adhesive and UV stabilizing properties. The acid resistant compounds as determined according to National Renewable Energy Laboratory standardized protocol NREL/TP-510-42618 (htps ://www. nrel.gov/docs/gen/ fy13/ 42618.pdf) comprise approx. 15-24 wt%, such as 18-24 wt% of the digestate dry matter, and the acid resistant compounds comprises 30-70% protein and protein complexes, 15-35% lignin and lignin derivatives; and 0.5-45% plastic derived compounds.

The protein fraction of the acid resistant compounds can be estimated from the nitrogen content corrected for ammonium nitrogen content is between 7 and 20 % by weight of the total digestate active filler composition, such as 10 - 17% or such as 12 - 15% by weight of the digestate active filler composition. The protein faction, however, is also comprised by the acid resistant compounds and represents 30-70% of the acid resistant compounds, such as 30-60%, 30-50%. 30-40% or such as 40-70%, 50-70% and 60-70%. The protein fraction is believed to be inert in asphalt mixtures. Nitrogen is present in asphaltenes and is thus a natural constituent in asphalt. The nitrogen in the digestate active filler might be incorporated in organic or inorganic complexes rather than present as protein.

According to C. Rodrigues and R. Hanumanthgari, The Shell Bitumen Handbook, 6 ^th ed., Chapter 8, p. 151-166 and G. Airey and A. Colop, ICE Manual of Construction Materials, Chapter 3, 2008, p 304 plastic derived compounds act as modifiers in asphalt mixtures. Plastic polymers are used to modify the properties of bitumen to optimize properties such as softening point, penetration, deformation resistance or viscosity. Plastic polymers can act as thermoelastic modifiers, elastomers or simply polymers. Ideally, bitumen stays soft at low temperatures to avoid cracking or fracturing and a hard and stiff bitumen at higher temperatures, but not so stiff as to be susceptible to cracking at low temperatures. Polymer modified bitumens can achieve these features compared to standard penetration grade bitumen products according to C. Rodrigues and R. Hanumanthgari, The Shell Bitumen Handbook, 6 ^th ed., Chapter 8, p. 151. The plastic compounds may be any of rubber, Polyurethane (Pll), Polyamide (PA), Polymethylmethacrylate (PMMA), Polycarbonate (PC), Polyvinylchloride (PVC), Polypropylene (PP), Polyethylene Terephthalate (PET), Polystyrene (PS) Polyethylene (PE), acrylonitrile butadiene styrene (ABS), polylactic acid (PLA), and polytetrafluoroethylene (PTFE). PE, PP, PVC and PS are thermoplastic polymers. PE, PP, PVC and PS are characterised by softening on heating and hardening on cooling and influences the penetration of the bitumen. Pll is a thermosetting polymer and PP acts a fibres according to C. Rodrigues and R. Hanumanthgari, The Shell Bitumen Handbook, 6 ^th ed., Chapter 8, p. 152. According to Cong et al., Construction and Building materials 225 (2019: p 1012-1025, Pll has been shown to modify asphalt mixtures towards improved temperature sensitivity so that the asphalt does not deform at high temperatures. Further, the rheology of foamed asphalt has been shown to be improved by Pll modification. Huang et al., Construction and Building Materials 356 (2022), shows that Pll also effects the water stability, fatigue resistance, anti-aging qualities and adhesion of asphalt whereas the temperature sensitivity does not seem to be proven according to this source. IOP Conf. Series: Materials Science and Engineering 203 (2017) 012004 claims that PET increases softening point of asphalt mixtures, improves stability, stiffness and viscosity, hence it can improve stripping, thermal cracking, temperature susceptibility, fatigue damage and rutting resistance.

The acid resistant compounds in the digestate active filler were analyzed and identified as disclosed herein. When measured as wt% of the total weight of the digestate active filler composition, the digestate active filler composition comprises in one embodiment:

Protein 5 - 13 wt% Lignin and lignin derivatives 3-7 wt%

Plastic derived compounds wherein 0.1 -0.8 wt% is visible plastic with a particle size of <1 mm and 0.001 - 0.007 wt% is microplastic with a particle size of 20-500 pm

Other acid resistant components 0.4 - 6.3 wt%

In a preferred embodiment, the acid resistant compounds in the digestate active filler comprises, when measured as wt% of the total weight of the digestate active filler composition:

Protein 7.4 - 11.1 wt%

Lignin and lignin derivatives 5 - 6 wt%

Plastic derived compounds wherein 0.2-0.7 wt% is visible plastic with a particle size of <1 mm and 0.001 - 0.005 wt% is microplastic with a particle size of 20-500 pm

Other acid resistant components 2.4 - 4.3 wt%

Another group of compounds that are defined herein to be comprised in the active filler is saturated and unsaturated long-chain fatty acids, which are apolar components such as palmitic, stearic and oleic acid, which has been quantified herein to amount to 0.1 - 5 wt%, such as 0.1-4 wt%, 0.1-3 wt%, 0.1-2 wt%, 0,1-1 wt% or such as 1-5 wt%, 2-5 wt%, 3-5 wt% and 4-5 wt%, or such as 0.2 wt% of the dry matter in a spot sample. These identified fatty acids have been found by J. Traffic Transp. Eng. (Engl. Ed.) 2022; 9 (2): 180e207, to act as rejuvenators in asphalt mixtures. The presence of saturated and unsaturated fatty acid compounds is known to have rejuvenating effects upon bitumen according to G. Airey and A. Colop, ICE Manual of Construction Materials, Chapter 3, 2008, p 304. Other apolar components found together with the identified fatty acids also have emulsifying properties according to M. Jair, The Shell Bitumen Handbook, 6 ^th ed., Chapter 9, p. 193-198, which is an advantage when working with cold mixed asphalt or foamed applications. J. Lu et al., Construction and Building Materials 391 (2023), 131735 shows that soybean oil has a softening effect on asphalt mixtures and acts as a rejuvenator.

The digestate active filler composition comprises between 18 and 24%, such as 18%, 19%, 20%, 21 %, 22% or 23% acid resistant compounds by weight of the digestate active filler composition.

The major fraction of the digestate active filler composition is ash, which accounts for 40 to 62%, such as 40 - 55% or 45 - 50% of ash by weight of the digestate active filler composition. This is the ash fraction obtained after a standard method by thermogravimetric analysis at 575°C. The ash fraction may comprise carbonate which is often used as an inert filler in asphalt compositions generally. In samples of the digestate active filler composition according to the invention it was found by thermogravimetric analysis at 900°C that the ash comprised up to 16% carbonate by weight of the ash composition treated by thermogravimetric analysis at 575°C. The ash fraction is known to contain compounds containing chemical compounds such as organo-zinc or calcium salts that can act as antioxidants according to C. Rodrigues and R. Hanumanthgari, The Shell Bitumen Handbook, 6 ^th ed., Chapter 8, p. 152, and some constituents of the ash fraction might act as pH agent and the ash fraction can thus act as chemical modifying agent through this effect.

The fraction identified as “other” is a fraction of various matter that cannot be identified as belonging to any of the other fractions. When analysing compositions derived from degradation of waste or other heterogenic material, it is common that some matter will remain unidentifiable. The fraction comprises compositions that is not inorganic residue after dry oxidation at 575°C, such as acid resistant components and non-degraded anhydrous carbohydrates.. This could be due to waxes or oil residues, or in-degradable protein complexes being underestimated.

In order to be qualified as a fine aggregate when the fine content is above 3%, the fines quality should be evaluated according to EN 933-9.

In order to be qualified as a filler in accordance with the European standard EN13043:2003, 100% of the particles should be 2 mm or less in size, 85 - 100% of the particles should 0.125 mm or less in size and 70 - 100% should be 0.063 mm or less in size.

In the digestate active filler composition presented herein, approximately all of the particles, that is at least 99% of the particles are 2 mm or less in size. Thus, at least 99%, such as 99.1 %, 99.2%, 99.3%, 99.4%, 99.5%, 99.6%, 99.7%, 99.8%, 99.9% or 100% of the particles are 2 mm or less in size. This can be found by passing the composition through a 2 mm sieve. The digestate active filler composition is obtainable from a method comprising subjecting MSW, to enzymatic and/or microbial treatment and anaerobic digestion and if no means for reducing the particle size is applied, the digestate active filler composition comprises on average 70% of particles with at size between 2 mm and 0.063 mm. A product can be added as a filler or as a combined fine aggregate and filler considering the particle size distribution. Alternatively, the product can be size reduced by any suitable means, for example milling to fill the demands for a filler. Even though size reduction such as milling is not required, it may be beneficial to get void of lumps, that is aggregates of filler composition because it is preferred that the digestate active filler composition is evenly applied to the asphalt composition. One mean for anti-lumping is drying of the digestate active filler composition and passing through a 2 mm sieve for separation of lumps but any means for anti-lumping available in the art may be applicable. In one embodiment, the digestate active filler composition is used in an asphalt composition without prior reduction of the particle size. In this embodiment, the composition is particularly useful as filler aggregate. Filler aggregate is of particular importance in the binder course of asphalt compositions and accordingly, for this embodiment the final application is suitably in asphalt binder course compositions.

In another embodiment, the digestate active filler composition is used in an asphalt composition after prior reduction of the particle size. In this embodiment, the composition is particularly useful as fine aggregate. Fines is of particular importance in the surface course of asphalt compositions and accordingly, for this embodiment the final application is suitably in asphalt surface compositions.

When the size distribution of the digestate active filler composition comprises a majority of particles between 2 mm - 0.063 mm, such as between 2 mm - 0.25 mm, or such as between 2 mm and 0.5 mm, or such as between 2 mm - 1 mm, the digestate active filler composition is preferably used in the binder course of asphalt compositions.

When the size distribution of the digestate active filler composition comprises a majority of particles between 0.063 mm - 0.03 mm or less, such as between 0.06 mm - 0.05 mm or less or such as between 0.042 mm - 0.03 mm or less or such as between 0.036 - 0.03 mm or less, or such as when all particles of the digestate active filler composition is below 0.03 mm, the digestate active filler composition is preferably used in the surface course of asphalt compositions

The digestate active filler composition tested herein comprises the particle size distribution as shown in Table A (un-milled sample) when no means for reducing the particle size was applied.

As shown herein, tests were also made with the digestate active filler composition wherein the particle size distribution in the digestate active filler composition was reduced to provide the following size distribution:

Table A: Particle size distribution for dried, loosened digestate

The particle size composition of digestate active filler composition can of course be adjusted in order to provide a certain share of particles within one or more desired size intervals depending on the intended use.

For digestate active filler compositions comprising from at least 99% to approximately 70% particles with particle sizes between 2 mm - 0.063 mm the following embodiments have been made.

In a preferred embodiment, at least 85% of the particles in the composition have a particle size of 0.25 mm - 2 mm.

In a preferred embodiment, at least 70% of the particles in the composition have a particle size of 0.5 mm - 2 mm.

For digestate active filler compositions comprising from 70% to approximately 100% particles with particle sizes of 0.063 mm - 0.001 mm the following embodiments are suggested.

In a preferred embodiment, at least 90% of the particles in the composition have a particle size of 0.086 mm or less.

In a preferred embodiment, at least 80% of the particles in the composition have a particle size of 0.06 mm or less.

In a preferred embodiment, at least 60% of the particles in the composition have a particle size of 0.03 mm or less.

In a preferred embodiment, 100% of the particles in the composition have a particle size below 0.06 mm.

In a preferred embodiment, 100% of the particles in the composition have a particle size below 0.06 mm and at least 60% of the particles in the composition have a particle size of 0.03 mm or less.

The active filler composition is preferably a digestate active filler composition, and the digestate may preferably be a result of AD treatment of a waste fraction, such as sewage sludge, seaweed, manure or household refuse such as MSW waste. Thus, in one embodiment the active filler is a digestate obtained from a method comprising: a. Subjecting a waste fraction to anaerobic digestion b. Separating the solid fraction from the liquid fraction of the digestate obtained in step a. c. Optionally subjecting the solid digestate fraction obtained in step b) to anti-lumping treatment and/or passing digestate through 2 mm sieve d. Optionally subjecting the digestate obtained in c) to drying and/or milling to reduce particle size

Preferably, the digestate active filler is obtained by a process further comprising two previous steps, i. Subjecting waste to enzymatic and/or microbial treatment ii. Subjecting the treated waste from step i) to one or more separation step(s) whereby a bioliquid fraction and a solid fraction is provided, wherein the bioliquid fraction obtained from step ii) is the waste fraction subjected to anaerobic digestion according to step a) in the method above.

The digestate active filler composition can be obtained from various waste treatment processes. Since the specific composition of a potential filler composition can be analysed by standard methods available in the art, the specific conditions applied for obtaining the digestate active filler composition is of less relevance.

However, in one aspect of the invention, the digestate active filler is obtained from a method comprising: i) Subjecting waste to enzymatic and/or microbial treatment ii) Subjecting the treated waste from step a) to one or more separation step(s) whereby a bioliquid fraction and a solid fraction is provided; and a) Subjecting the bioliquid fraction to anaerobic digestion b) Separating the solid fraction from the liquid fraction of the digestate obtained in step a) c) Optionally subject the solid digestate fraction obtained in step b) to anti-lumping treatment and/or passing digestate through 2 mm sieve d) Optionally subjecting the digestate obtained in c) to drying and/or milling to reduce particle size. The method can be performed within a single waste processing plant comprising one or more bioreactors and/or one or more downstream AD reactors or the waste treatment process providing the digestate active filler composition can be performed at two or more different and possibly independent waste processing and/or biogas production sites.

Step i), ii) and a) in the method are commonly applied steps in waste treatment methods, and details regarding one or more of these steps have been disclosed in for example WO2014/198274, WO2013/18778, WO 2022/096406 and WO 2022/096517.

The steps of the method can be described as follows:

The enzymatic and/or microbial treatment of the waste in step i) can for instance be performed in a bioreactor. The treatment is performed by adding one or more enzymes and by the bacteria present in the waste. Optionally, standard, cultivated, or manipulated yeast, bacteria, or any other microorganism capable of converting the organic matter present in the waste into compositions suitable for subsequent biogas production in an anaerobic digestion process may be added to the bioreactor. The enzymes are supplied in either native form or in form of microbial organisms expressing the enzymes.

The enzymatic and/or microbial treatment in step i) may be performed by adding one or more enzymes, supplied in either native form and/or in form of microbial organisms giving rise to the expression of such enzymes; and/or by the bacteria present in the waste and/or optionally by adding standard, cultivated, or manipulated yeast, bacteria, or any other microorganism capable of converting the organic matter present in the waste into organic acids or other compositions, such as lactic acid, 3-hydroxypropionic acid (3-HPA), 1 ,4-butanediol (BDO), butanedioic acid (succinic acid), ethane-1 ,2-diol (ethylene glycol), butanol or 1 ,2-propanediol (propylene glycol), suitable for subsequent biogas production in an anaerobic digestion process.

Microorganisms that may be added to the bioreactor in step i) include yeasts, and/or fungi and/or bacteria.

Other microorganisms that may be added to the bioreactor in step i) include bacteria that can efficiently ferment hexose and pentose including but not limited to cellobiose, glucose, xylose and arabinose to short chain organic acids including but not limited to citric acid, lactic, formic acid, acetic acid, butyric acid, valeric acid, isovaleric acid and propionic acid as well as alcohols including but not limited to ethanol. The fermenting microorganisms may have been genetically modified to provide the ability to ferment pentose sugars, such as xylose utilizing, arabinose utilizing, and xylose and arabinose co-utilizing microorganisms.

The fermenting organisms may comprise one or more polynucleotides encoding one or more cellulolytic enzymes, hemicellulolytic enzymes, and accessory enzymes described herein.

The microorganisms present in the waste or added to the bioreactor, may produce fermentable sugars and organic acid or other compositions, such as lactic acid, 3-hydroxypropionic acid (3-HPA), 1 ,4-butanediol (BDO), butanedioic acid (succinic acid), ethane-1 ,2-diol (ethylene glycol), butanol or 1 ,2-propanediol (propylene glycol), that may be used as feed in a subsequent anaerobic digestion process. These organic acids or other compositions further include acetate, propionate and butyrate. Waste that is suitable for treatment normally comprises, at least, lactic acid producing bacteria.

The treatment in step i) may comprise addition of cellulase activity by inoculation with one or more microorganism(s) that exhibits extracellular cellulase activity.

In step i) the waste may also be treated with an enzyme composition wherein the enzymes are added to the waste independently from the enzymes present within the microorganisms already present or added to the waste. Suitable enzyme compositions are well known in the art and are commercially available.

Suitable enzyme blends are cellulolytic background composition (CBC) comprising a commercial cellulolytic enzyme preparation. Examples of commercial cellulolytic enzyme preparations suitable for use in the method according to the present invention include but is not limited to, for example, CELLIC® CTec (Novozymes A/S), CELLIC® Ctec2 (Novozymes A/S), CELLIC® Ctec3 (Novozymes A/S), CELLUCLAST® (Novozymes A/S), NOVOZYM™ 188 (Novozymes A/S), SPEZYME™ CP (Genencor I nt.), ACCELLERASE™ TRIO (DuPont), FILTRASE® NL (DSM); METHAPLUS® S/L 100 (DSM), ROHAMENT™ 7069 W (Rohm GmbH), or ALTERNAFUEL® CMAX3™ (Dyadic International, Inc.).

In addition to the CBC, further enzyme activity may be added from individual sources or together as part of enzyme blends. Suitable blends include but are not limited to the commercially available enzyme compositions Cellulase PLUS, Xylanase PLUS, BrewZyme LP, FibreZyme G200 and NCE BG PLUS from Dyadic International (Jupiter, FL, USA) or Optimash BG from Genencor (Rochester, NY, USA). For treatment of MSW suitable CBCs comprises enzymatic activity in accordance with the activity of ACCELLERASE® TRIO™ (Genencor Int.), Cellic Ctec2 (Novozymes A/S) or Cellic Ctec3 (Novozymes A/S) or Cellic Ctec3 (Novozymes A/S).

The enzymatic treatment of the biodegradable parts of the waste concurrently with microbial fermentation according to step i) may be performed at a temperature above 20°C and up to 75°C resulting in liquefaction and/or saccharification of biodegradable parts of the waste and accumulation of sugars and other soluble degradation products.

The waste, e.g., MSW, may have a Dry Matter (DM) content in the range 10%-90%; 20%-85%; 30%- 80%; 40%-75%; 50%-70%; or 55%-65 % (wt%); and/or around 10%; 15%; 20%; 25%; 30%; 35%; 40%; 45%; 50%; 55%; 60%; 65%; 70%; 75%; 80%; 85%; or 90% (wt%). Preferably, the DM content of the MSW is in the range of 10 - 80%, such as 20 - 70%, 30 - 60% and 20 - 50%.

In order for the enzymatic and/or microbial liquefaction of the waste in the bioreactor in step i) to provide a bioliquid comprising an optimum amount of short chain carboxylic acids and sugars such as glucose, xylose, arabinose, lactic acid/lactate, acetic acid/acetate and/or ethanol, the pH in the bioreactor should generally remain within a pH range of between pH 2 - 6.5.

Step ii) is a separation step where the bioliquid is separated from the non-degradable solid waste fractions. Clean and efficient use of the degradable component of waste, such as MSW, combined with recycling typically requires some method of sorting or separation to separate degradable from non-degradable material. The separation in step ii) may be performed by any means known in art, such as in a ballistic separator, washing drums and/or hydraulic presses. In one embodiment the separation is performed before the enzymatic treatment. Separation of liquid and solids can e.g. be done in different presses (such as screw and/or piston presses) or e.g. using a simpler sieve function. A ballistic separator is typically used to separate the solids into fractions and only secondarily a liquid separation.

In step a), the bioliquid fraction obtained in step ii) is subjected to anaerobic digestion. Anaerobic digestion (AD) is a series of biological processes in which microorganisms break down biodegradable material in the absence of oxygen. One of the end products is biogas, which can be combusted to generate electricity and/or heat, or can be processed into renewable natural, biomethane gas and/or transportation fuels. A range of anaerobic digestion technologies exists in the state of the art for converting waste, such as MSW, municipal wastewater solids, food waste, high strength industrial wastewater and residuals, fats, oils and grease (FOG), and various other organic waste streams into biogas. Many different anaerobic digester systems are commercially available, and the skilled person will be familiar with how to apply and optimize the anaerobic digestions process. The metabolic dynamics of microbial communities engaged in anaerobic digestion are complex.

In typical anaerobic digestion (AD) for production of methane biogas, biological processes mediated by microorganisms achieve four primary steps - hydrolysis of biological macromolecules into constituent monomers or other metabolites; acidogenesis, whereby short chain hydrocarbon acids and alcohols are produced; acetogenesis, whereby available nutrients are catabolized to acetic acid, hydrogen and carbon dioxide; and methanogenesis, whereby acetic acid and hydrogen are catabolized by specialized archaea to methane and carbon dioxide. The hydrolysis step is typically rate-limiting and dependent on the biomass type. In the bioliquid it is the methanogens that limits the processing rate. From AD is furthermore obtained digestate, comprising a solid fraction and a liquid fraction (reject water), in particular comprising a water-like liquid with separable suspended particles. Such solid digestate is, in one embodiment of the present invention, the digestate active filler composition according to the invention.

The anaerobic digestion step a) may comprise one or more reactors operated under controlled aeration conditions, eliminating or minimizing the available oxygen, in which methane gas is produced in each of the reactors comprising the system. The AD reactor(s) can, but need not, be part of the same waste processing plant as the bioreactor in step i) and can, but need not, be connected to the bioreactor in step i). Moreover, the AD process may be in the form of a fixed filter system. A fixed filter anaerobic digestion system is a system in which an anaerobic digestion consortium is immobilized, optionally within a biofilm, on a physical support matrix.

In step b) the digestate obtained from step a) is separated into a liquid phase and a solid digestate and the solid digestate is optionally subjected to step c).

The digestate comprises both solids and liquids and these fractions may be used for various purposes. The solid-liquid separations can for instance be done by decantation, centrifugation and/or sedimentation. Usually, the anaerobic digestate comprises mainly water, wherein the digestate has a total solid content of about 4-8% and after the dewatering the solid digestate has a total solid content of 25-45% by weight, the rest being water. It also comprises non-degradable organics, suspended solids and dissolved matter such as dissociated salts and has alkaline pH.

After separation of the liquid phase, the solid digestate fraction obtained in step b) is optionally subjected to anti-lumping treatment and/or passing digestate through 2 mm sieve. The purpose of this step is to avoid lumping of the digestate. The particles tend to aggregate to each other forming lumps of various sizes. When the digestate is used in asphalt compositions as digestate active filler it is, however, beneficial that the filler composition is added and distributed evenly in(to) the asphalt composition. Therefore, an anti-lumping step may be required. Anti-lumping can be performed using any suitable means and methods known in the art. One such method is normally obtained when drying the solid digestate until constant mass. This is normally obtained when the solid digestate has between 0-20% moisture, such as 0%, 2.5%, 5%, 7.5%, 10% 12.5%, 15%, 17.5% or 20%. Preferably, the solid digestate is dried until having between 0-10% moisture, such as 5% or such as 0%, 2%, 4%, 6%, 8% or 10%. The solid digestate can be dried by subjecting to temperatures of up to about 105°C, such as 70°C, 75°C, 80°C, 85°C, 90°C, 95°C, 100°C or 105°C for a period of time sufficient to obtain constant mass. Normally, the time required is from 20 to 30 hours, such as 24 hours.

Step d) is an optional step that is to be selected when particles of smaller sizes are desirable for the specific use of the MSW derived anaerobic digestate active filler composition. If step d) is selected, the solid digestate obtained from step c) is subjected to drying and/or milling to reduce particle size. Depending on the milling method applied to reduce particle size a previous drying step may be required. Some milling means can only be applied on dry material whereas other means for milling provides sufficient particle size reduction when applied on wet material. The intensity of the milling can be adjusted, for example by adjusting the duration of time of subjecting the solid digestate and thereby provide various particle size fractions.

The enzymatic and/or microbial waste treatment process described in step i) is applicable to a wide range of waste comprising organic matter. Agricultural material/waste, household waste and municipal waste are examples of sources containing a high content of dry matter and a certain content of organic material. Since it is the organic fraction of waste subjected to enzymatic and/or microbial treatment in a bioreactor in step i) that provide a bioliquid substrate suitable for the anaerobic digestion in step a), waste comprising organic matter is a suitable feed. Examples of suitable waste includes MSW, agriculture waste, industrial waste, waste fractions from recycling facilities, and garden refuse. The process is applicable to unsorted as well as to sorted waste. In preferred embodiments, the waste is sorted or unsorted MSW.

The digestate active filler composition according to the invention can also be obtained from other waste treatment processes that are different from the method described above. Regardless of the method applied to provide the digestate active filler composition, MSW is a preferred waste source due to its composition of mixed organic waste fractions. It is shown herein that the digestate active filler composition comprises very little variation in the amount of each component when obtained from different sources and at different points in time over the year. Thus, the composition obtained appears very robust and it is therefore believed that the same composition of ingredients and their amounts, will be in line with the digestate active filler composition disclosed herein regardless of the specific organic waste it is obtained from.

In view of practical handling and storage of the digestate active filler mixture composition as well as in view of its use in asphalt compositions, it is preferred that the digestate active filler composition is dried until constant mass before use due to the mechanical strength of the asphalt risk being compromised if course and fine aggregates and filler and other additives cannot be distributed evenly due to lumping.

As disclosed in the examples herein, the digestate active filler composition was added to an asphalt surface composition and provided an impact on the softening of the asphalt surface composition by reducing the softening point temperature of the asphalt surface composition. The addition of the digestate active filler composition to an asphalt surface composition moreover provided an antiageing effect. Moreover, the addition of the digestate active filler composition to an asphalt surface composition provided a rejuvenating effect to the asphalt surface composition. Without being bound by theory, it is our theory that the interaction between the digestate active filler and the bitumen binder causing modification of the bitumen properties, e.g. the softening, the anti-ageing, and rejuvenating effect observed in the asphalt compositions tested, is derived from a mixture of effects. This is most likely caused by e.g., the apolar components such as saturated and unsaturated fatty acids, microplastic, inorganic complexes such as calcium salts, fibres such as lignin and cellulose and the chemical effects of a high pH in the material.

Thus, one aspect of the present invention provides the use of the digestate active filler composition according to the invention in the binder course and/or in the surface course of asphalt compositions.

It is shown here that asphalt surface compositions comprising quite a large amount of the digestate active filler composition the invention showed increased softening, anti-ageing and rejuvenation while upholding sufficient tensile strength.

In a preferred embodiment, the digestate active filler composition, such as the digestate active filler of the invention is used in the binder course and/or in the surface course of asphalt compositions wherein 60% by weight of the total fine aggregate components in the binder course and/or in the surface course is a digestate active filler composition according to the invention. Accordingly, such use include use wherein 10 - 60%, 15 - 55%, 20 - 50%, 25 - 45%, or 30 - 40% by weight of the total fine aggregate components in the binder course and/or in the surface course is a digestate active filler composition according to the invention. In another preferred embodiment, the digestate active filler composition of the invention is used in the binder course and/or in the surface course of asphalt compositions wherein 60% by weight of the total filler aggregate components in the binder course and/or in the surface course is a digestate active filler composition according to the invention. Accordingly, such use include use wherein 10 - 60%, 15 - 55%, 20 - 50%, 25 - 45%, or 30 - 40% by weight of the total filler aggregate components in the binder course and/or in the surface course is a digestate active filler composition according to the invention.

In a preferred embodiment, the present invention provides use of the digestate active filler composition according to the invention for one or more of softening, anti-ageing and rejuvenation of the binder course and/or the surface course of an asphalt composition. Apolar components such as fatty acids are known to rejuvenate bitumen. Several polymers are known to soften bitumen and salts such as calcium salts are known to protect bitumen from oxidation, as is an increase in pH. All these components are present in the digestate active filler.

The main difference between the surface composition and the binder composition of an asphalt of relevance to the use of the digestate active filler composition is the particle size of the aggregates in the asphalt course.

Since the aggregates in the surface course have smaller particle sizes, use of digestate active filler compositions of the invention wherein the particle size composition has an overweight of smaller particles are preferred in the surface course.

Since the aggregates in the binder course have larger particle sizes, use of digestate active filler compositions of the invention wherein the particle size composition has an overweight of larger particles are preferred in the binder course.

When used in asphalt surface and/or asphalt binder compositions, the digestate active filler composition of the present invention may represent up to 100%, such as 60% by weight of the total filler aggregate components in the binder course and/or in the surface course.

When used in asphalt surface and/or asphalt binder compositions, the digestate active filler composition of the present invention may represent up to 100%, such as 60% by weight of the total fine aggregate components in the binder course and/or in the surface course.

In one aspect, an asphalt binder course composition comprising up to 100% of digestate active filler composition according to the invention of the total amount by weight of fine aggregate components in the binder course composition is provided. This includes asphalt binder course compositions wherein, 1-80%, 5-70%, 10-60%, 15 - 55%, 20 - 50%, 25 - 45%, 30 - 40%, or 1-10% by weight of the total filler components in the binder course is a digestate active filler composition according to the invention.

In a preferred embodiment of this aspect, the digestate active filler composition according to the invention is a composition wherein 0 - 30% of the particles in the composition have a particle size of 0.063 mm or less.

In another aspect, an asphalt surface course composition comprising up to 100% of digestate active filler composition according to the invention of the total amount by weight of filler aggregate components in the surface course composition is provided. This includes asphalt surface course compositions wherein, 1-80%, 5-70%, 10-60%, 15 - 55%, 20 - 50%, 25 - 45%, 30 - 40%, or 1-10% by weight of the total filler aggregate components in the surface course is a digestate active filler composition according to the invention.

In a preferred embodiment of this aspect, the digestate active filler composition according to the invention is a composition wherein 100% - 70% of the particles in the composition have a particle size of 0.063 mm or less.

In another aspect, an asphalt composition comprising up to 100% of digestate active filler composition according to the invention of the total amount by weight of fine aggregate components or filler aggregate components in the surface course and in the binder course is provided. This includes asphalt compositions wherein, 1-80%, 5-70%, 10-60%, 15 - 55%, 20 - 50%, 25 - 45%, 30 - 40%, or 1-10% by weight of the total fine aggregate components or of the total filler aggregate components in the surface course and in the binder course is a digestate active filler composition according to the invention.

EXAMPLES

Example 1 Preparation of composition and size distribution

1.2 Preparation of digestate active filler composition

Samples of starting material for the digestate active filler composition was obtained from a waste treatment process wherein MSW was subjected to enzymatic liquefaction, and the liquid fraction thus obtained was subjected to anaerobic digestion.

The first steps applied for preparing the samples were made as described in WO 2020/002153 A1 Example - Preparation of Digestate additives point 1.1 step 1 , step 2, and 3 three (page 65 line 23 to page 67 line 24) hereby incorporated by reference. The produced bioliquid was used for biomethane production in a full-scale anaerobic digester. The anaerobic digesters were four 4.300 cubic meter liquid filled tanks (0 31 x 8 m) where the biological conversion took place. The tanks were equipped with four agitators for each tank. The effluent was discharged by bottom extraction to a hygienisation system where the digestate was heated to 70°C for 1 hour. The hygienised digestate was dewatered with decanter centrifuges to produce the solid digestate with a suspended dry matter of 25-40 wt%. A cationic polyacrylamide flocculant was used to aid the de-watering process. The concentrated digestate from anaerobic digestion periods with stable biogas production was collected in 500L pallet tanks and transported to the Renescience laboratory in Denmark by currier.

The 300-400 kg monthly spot samples were subjected to drying in a EMMERT UF 750 convectional oven at 105 °C for 24 hours. The material was loosened by running it through an Retsch SM300 mill with a 2.0 mm screen inserted. The product was - in some experiments - subsequently milled in a Retsch planetary ball mill PM 100 to reduce the particle size further.

Samples were collected monthly in August 2021 (Sample 1), September 2021 (Sample 2), October 2021 (Sample 3), November 2021 (Sample 4) December 2021 (Sample 5), January 2022 (Sample 6), February 2022 (Sample 7), March 2022 (Sample 8), April 2022 (Sample 9), May 2022 (Sample 10), July 2022 (Sample 11) and August 2022 (Sample 12).

1.2 Size distribution determination

All digestate active filler samples were analysed by laser diffraction before and after milling to determine the size fractions of the composition on a HELOS/KF Particle Size Analyser at 1.5 bar with vibrational in feeder with 65% feed rate before and after milling in order to determine the particle size fractions of the composition.

Table 1 discloses the percentage of composition passing through various sieve sizes after milling from sample Samples from 3, 4, and 5 were additionally subjected to chemical composition analysis. Table 1: Size fractions of the particles in the filler composition

The particle size distribution for milled and unmilled digestate is seen from Figure 1 A and B, respectively. Example 2 Composition analysis of digestate active filler composition.

The samples of digestate active filler described in Example 1 were subjected to chemical compositional analysis, and DCM extraction for analysis of the composition. 2.1 Chemical composition analysis

Test method: Sluiter et al 2008 (NREL/TP-510-42620): Preparation of Samples for Compositional Analysis, Sluiter et al 2008 (NREL/TP-510-42619): Determination of Extractives in Biomass and Sluiter et al 2008 (NREL/TP-510-42618): Determination of Structural Carbohydrates and Lignin in Biomass adapted to use a Dionex Ultimate 3000 HPLC system equipped with a Rezex Monosaccharide H+ column from Phenomenex.

Table 2 discloses the extractives which was extracted prior to analysis of the chemical composition of solid digestate obtained in Sample 1 , Sample 2, and Sample 3, respectively.

Table 2: Extractives in digestate samples according to NREL/TP-510-42619: Determination of Extractives in Biomass

EXTRACTIVES (WT%) ol/toluen Ethanol Water total 3.4 0.3 5.5 9.2 2.4 0.2 4.5 7.1 | 3.2 0.3 5.5 9.0

Extraction prior to analysis of anhydrous carbohydrates and acid resistant components, assumed to be Klason-lignin when analysing lignocellulosic materials, is performed to prevent extractives causing poor reproducibility and/or interfering with determination of Klason-lignin. Thus, the analysis was performed with and without extraction for three samples and it was concluded that the extractables in solid digestate does not interfere with the analysis (data not shown). The remaining samples was accordingly analysed without prior extraction.

Table 3 discloses the anhydrous carbohydrates detected in the chemical composition of dried solid digestate samples obtained in sample 1 , sample 2, sample 3, sample 4, sample 5, sample 6, sample 7, sample 8, sample 9, sample 10, sample 11 and sample 12 respectively. Standard deviations for duplicate determination are given for sum of anhydrous carbohydrates.

Table 3: Anhydro carbohydrates (glucan, xylan and arabinan) in digestate samples analysed according to NREL/TP-510-42618

Table 4 discloses the acid resistant content detected in the chemical composition of dried solid digestate samples obtained in sample 1 , sample 2, sample 3, sample 4, sample 5, sample 6, sample 7, sample 8, sample 9, sample 10, sample 11 and sample 12 respectively. Standard deviations for duplicate determination are given for sum of acid resistant components. The acid resistant components are usually described as Klason-lignin when performing the analysis on lignocellulosic materials where the acid resistant components can be assumed to be lignin. The assumption is not assumed to be valid for solid digestate as the material is not a pure lignocellulosic material. Samples with an ash content above 10 wt% will likely cause overestimation of the Klason-lignin content due to neutralisation of the acid used, this is most likely the case for the digestate active filler composition. Wax or oils can also cause positive interference on the analysis, however no difference with and without extraction was observed. Further, samples containing protein will cause overestimation of the acid resistant components and the protein content estimated from the nitrogen content should be subtracted the value. Protein is estimated for the samples, see Example 2.2, Table 6. The acid resistant components are corrected for the estimated protein amount. The corrected and uncorrected amount of acid resistant components are seen from Table 4.

Table 4: Acid resistant components determined according to NREL/TP-510-42618 with and without correction for estimated protein content from nitrogen content

Table 5 discloses the inorganic ash fraction detected in the chemical composition of dried solid digestate samples obtained in sample 1 , sample 2, sample 3, sample 4, sample 5, sample 6, sample

7, sample 8, sample 9, sample 10, sample 11 and sample 12, respectively. Standard deviations for duplicate determination are given for sum of anhydrous carbohydrates.

Table 5: Ash fraction determined according to NREL/TP-510-42618 At the Northwich Renescience plant, the solid digestate is sampled routinously every workday. 50 g of sample is sampled and stored in a plastic container in a refrigerator at4°C. Samples are compiled over approximately two weeks until the container is full and contains approximately 500 g of sample.

The combined sample representing the average of two weeks production of solid digestate was analyzed by placing the material in a muffle furnace at 550°C until constant weight and calculating the loss on ignition and the ash content. It was found that the ashsso’c content on average was 55±8 wt% of the dry matter of the solid digestate for 37 routine samples taken in the period from January 2021 to December 2022. The variation observed in the data set was from 34 to 60 wt% ashsstrc of the dry matter of solid digestate.

The chemical composition analyses showed that the digestate active filler composition had a high ash content and a substantial protein content. Further, a substantial amount of carbohydrates, glucan, xylan and arabinan, was identified as well as a substantial fraction of acid resistant components was identified.

2.2 Protein estimation

The acid resistant components fraction comprises proteins, as estimated in example 2.1.

Protein is estimated by analysing the dried solid digestate samples for total nitrogen content and estimating the protein content. The nitrogen content was determined according to the DUMAS technique, AOAC Official Methods of Analysis (1990), Method 949.12, where the dried sample is totally combusted in an oxygen enriched atmosphere in a reaction tube. The nitrogen and carbon are carried by a constant flow of carrier gas (helium) through an oxidation catalyst, and then through reduced copper wires, where excess oxygen is removed, and nitrogen oxides are reduced to elemental nitrogen. The nitrogen and carbon products are separated through a chromatographic column. As the products are eluted from this column, they pass through a Thermal Conductivity Detector, which generates and electric signal proportional to the amount of nitrogen and carbon present.

The determination of Nitrate-N and Nitrite-N is based on the formation of a diazo compound between nitrite and sulphanilamide. This compound is then coupled with N-1-Napthylethylenediamine dihydrochloride to give a red azo dye. The color is measured at 540nm. In channel one, nitrate is reduced quantitatively to nitrite by cadmium metal in the form of an open tubular cadmium reactor (OTCR). The nitrite and reduced nitrate are therefore both measured as total oxidized nitrogen. In channel two, nitrite is measured. Nitrate-N is determined by deducting the nitrite figure from the total oxidized nitrogen. In channel three, ammonium reacts with alkaline hypochlorite and phenol to form indophenol blue. Sodium nitroprusside acts as a catalyst in formation of indophenol blue which is measured at 640nm. Precipitation of calcium and magnesium hydroxides is eliminated by the addition of a combined potassium sodium tartrate/sodium citrate complexing reagent.

The protein content is calculated by subtracting the ammonium nitrate and the nitrite, which is not detected above 10 mg per kg dry matter in the digestate samples and thus ignored, from the total nitrogen and multiplying the result with a factor of 6.25. The results from tested samples 1 to 9 are shown in table 6. This estimation is done accordance with J. Liebetrau and D. Pfeiffer, Biomass energy use, Vol. 7, collections of Methods for Biogas in biogas digestates.

Table 6: Estimated protein content from nitrogen content (DUMAS method) and ammonium nitrogen and nitrate (not shown, always below 10 mg per kg dry matter)

At the Northwich Renescience plant, the solid digestate was sampled routinously every workday. 50 g of sample is sampled and stored in a plastic container in a refrigerator at 4°C. Samples are compiled over approximately two weeks until the container is full and contains approximately 500 g of sample.

The combined sample representing the average of two weeks production of solid digestate was analyzed according to the DUMAS method described above. It was found that the estimated protein content on average was 11 ±4 wt% of the dry matter of the solid digestate for 37 routine samples taken in the period from January 2021 to December 2022. The variation observed in the data set was from 5 to 17 wt% estimated protein of the dry matter of solid digestate. The variation over time is seen from Figure 9. Example 3 Mixing the aggregate(s), filler(s) and binder(s) with digestate additive to obtain an asphalt mixture composition

A standard sample of asphalt mixture was prepared. The recipe used was to produce an AC surf 11 top layer asphalt as shown in Table 7.

Table 7: Asphalt recipe for samples with 6.2% 70/100 in an AC surf composition with limestone filler

Every month, a sample of asphalt mixture was prepared. The recipe used was to produce an AC11 surf asphalt sample. For the tests the filler composition obtained from Examples 1 and 2 was added from the respective month tested by replacing 50% of the filler with digestate active filler composition of the invention to provide a top layer asphalt composition with particle sizes and wt% of the components as shown in Table 8. Table 8: Asphalt recipe for samples with 6.2% 70/100 in an AC surf composition with limestone filler, where 50% is replaced with ball-milled digestate

Before the mixing begins, mixing containers were pre-heated to 155 ± 25 °C. Then aggregates preheated to 155 ± 25 °C, were weighed into the containers. Aggregates were mixed. Bitumen used in this assay, a 70/100 penetration grade bitumen, was then added until reaching the desired binder content in the asphalt mixture composition. Filler was then added. All components were then thoroughly mixed for approximately 3-5 minutes and continuously mixed in said containers, until good mixing is acquired, where the aggregates was entirely coated with the binder. The mixture was poured into a suitable form and manually dispersed before it was compacted. The asphalt was left to cool to 90-100°C, measured by an I R handheld thermometer, before the final compacting. The mixture was then allowed to cool to room temperature over night, before the slab is unmolded. The sample was stored in a climate chamber at 15°C to allow the binder(s) to harden before preparing test specimens.

To evaluate if particle size reduction can be omitted and the digestate active filler can be used as a mixed filler consisting of both fines and fine aggregates (sand equivalents), a composition was compared with un-milled solid digestate replacing 50% of filler material with the digestate active filler material.

Table 9: Asphalt recipe for samples with 6.2% 70/100 in an AC surf composition with limestone filler, where 50% is replaced with digestate. Further, some fine aggregates (sand equivalents) were replaced.

A series of recipes with a harder, 40/60 grade, bitumen was also conducted. A reference asphalt sample was produced, the recipe is shown in Table 10. Table 10: Asphalt recipe for samples with 5.8% 40/60 in an AC surf composition with limestone filler

For the tests the filler composition obtained from Examples 1 and 2 was added from the October (3), January (6) and April sample (9) tested by replacing 50% of the filler with unground digestate active filler composition of the invention to provide a surface layer asphalt composition with particle sizes and weight% of the components as shown in Table 11.

Table 11 : Asphalt recipe for samples with 5.8% 40/60 bitumen in an AC surf composition with limestone filler, where 50% is replaced with digestate active filler. Further, some fine aggregates (sand equivalents) were replaced.

Example 4 Penetration and softening point of bitumen extracted from fresh and aged asphalt composition comprising the filler composition provided in Examples 1 and 2

Penetration and softening point of bitumen extracted from an asphalt composition according to Example 3, Table 7 and Table 8, comprising the digestate active filler composition described in Examples 1 and 2 were tested.

From samples of digestate taken at the Renescience Northwich plant every month for a year, preferably between the 8 ^th and the 15 ^th of each month for a year, a 25 kg batch of the above asphalt composition (AC 11 Surf) was made with aggregates, 70/100 bitumen and ball-milled digestate. The test was performed within 1.5 hours after filling the molds with liquid bitumen for both virgin and aged test samples. The aging was performed by leaving a sample for 3 weeks aging in a closed internal ventilated oven at 85°C. This corresponds to approximately 6 years of service life. The aging can be made by other commonly applied aging methods, such as the Rolling Thin Film Oven Test (RTFOT), temperature ageing and the Pressurised Ageing Vessel (PA ) test. The bitumen from the fresh and aged samples was extracted with dichloromethane prior to analysis according to EN 12697-1 :2020 soxhlet extractor method.

The penetration tests were performed using the needle penetration test (Figure 2) in accordance with EN 1426:2015. The penetration is expressed as the distance in tenths of a millimetre that a standard needle will penetrate vertically into a sample of the material under specified conditions of temperature, load and loading duration.

The softening point tests were performed using the Ring and Ball test (Figure 3) in accordance with EN 1427:2015. The softening point is defined as the mean of the temperatures at which two discs of bituminous binders, cast in shouldered brass rings, heated at a controlled rate in a liquid bath while each support a brass ring, softens enough to allow each ball to fall a distance of 25.0±0.4 mm. Figure 4 shows needle penetration (A), (B) softening point, and (C) penetration index, respectively, in aged and fresh samples for a reference AC11 surf sample with 6.2% 70/100 bitumen and 12 samples of AC11 surf with 6.2% 70/100 bitumen with 50% filler (limestone: Wigras 40) replacement with digestate (Sample 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 and 12). As shown in Figure 4 these tests showed that penetration of the binders extracted from the asphalt composition comprising the digestate active filler composition are softer than the penetration of binders extracted from the reference. The softening point numbers are matching with penetration expressed in the Penetration Index (PI) as described in The Shell Bitumen Handbook by Hunter, Self and Read (2003) and therefore meets the requirement of value of -1.5 to 0.7. ring and ball) [ C] + 500 ■ log(P ₂ 5°c) ^— 1952 ing and ball) [°C] - 50 ■ log(P ₂₅ = _c ) + 120

Aged samples will have a higher PI whereas the softening points in the aged samples are slightly higher than in the reference samples. The softening points in all samples were above the minimum value of -1.5.

Before aging, the penetration index was found to be on average -0.9 for the reference sample and - 1.2 for the samples comprising digestate active filler. This difference between the reference asphalt samples and the 7 samples with 7 different spot samples of dried solid digestate replacing 50% of the filler material is not statistically significant (P=0.10). After aging, the mean values were found to be significantly lower for the asphalt samples containing solid digestate compared to the reference sample, meaning that the asphalt is softer (P=0.03 with R ² explaining 65% of the variation).

This shows that there is a positive effect of solid digestate replacing the filler in asphalt, as the penetration index of the new asphalt samples is not significantly different, thus the asphalt remains strong, but the digestate seems to counteract the aging effect with the asphalt becoming stiff and brittle as was observed on the reference sample.

The below Table 12 shows the results obtained in the penetration and softening point tests for the reference sample and for the bitumen samples 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 and 12, respectively. The analysis was conducted shortly (within 1.5 hours) after addition of the filler composition and after ageing.

Table 12: Needle penetration (EN 1426:2015) and softening point (EN 1427:2015) for bitumen extracted from fresh and aged asphalt samples, reference sample and sample 1 to 12 with digestate replacing 50% filler

Figure 4 shows the penetration (a), softening point (b), and penetration index (specification limit: - 1.5) (c) of reference asphalt samples compared with samples with 50% filler replacement with solid digestate. Example 5 Indirect Tensile Strength (ratio) of asphalt compositions provided in Example 3 comprising the digestate active filler composition as described in Example 1 and 2

The asphalt compositions described in Example 3, Table 7 and Table 8, comprising the digestate active filler described in Examples 1 and 2 in an AC11 surf with 6.2% 70/100 bitumen were subjected to test for indirect tensile strength according to EN 12697-23:2017. Every month a 25 kg batch of the above asphalt composition (AC 11 Surf) was made with 70/100 bitumen. The test was performed within 42 days after production of the test specimens. Three produced test specimens were cut from the produced asphalt slap and analysed as is and the indirect tensile strength of the dry specimens is recorded, ITSdry. Three test specimens were soaked in water at 40°C for 72 hours and the indirect tensile strength was recorded for these specimens, ITSwet. The indirect tensile strength ratio is calculated as ITSwet relative to ITSdry.

Table 13: Indirect tensile strength before (ITSdry) and after wetting (ITSwet) and indirect tensile strength ratio (ITSR) for asphalt samples; reference sample with limestone filler and sample 1 to 12 with digestate replacing 50% filler in triplicate for each sample

A graphical visualisation is seen from Figure 5. Figure 5 shows indirect tensile strength for wet and dry samples for a reference AC11 surf sample with 6.2% 70/100 bitumen and 12 samples of AC11 surf with 6.2% 70/100 bitumen with 50% filler (limestone: Wigras 40) replacement with digestate (Sample 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 and 12). In Figure A, the indirect tensile strength of the dry and wet specimen is compared in a box and whiskers plot for the reference and samples with replacement of filler with digestate. In Figure 5 B, the ITSR is compared in a box and whiskers plot. In Figure 5 C and D are the ITS and ITSR respectively plotted as a function of sampling date. The tensile strength of the samples is comparable to the control sample and within commonly accepted standard. The indirect tensile strength ration is not significantly lower for the samples tested compared to the reference and higher than the 80% required by the industry (Shell bitumen handbook, 6 ^th ed., page 456).

Example 6 Tri-Axial rutting tests

The asphalt compositions described in Example 3, Table 7 and Table 8, comprising the digestate active filler described in Examples 1 and 2 and a reference sample without digestate were subjected to Tri-Axial rutting tests according to EN 12697-25:2016, method B. Three test specimens with a height of 60±2 mm and a diameter of 148±5 mm was cut from each of the 12 asphalt slaps containing digestate and the reference asphalt. The test was conducted within 14 to 42 days after producing the asphalt sample and the sample specimen was stored in a climate chamber at approximately 15°C until testing. The test specimen was cured to ambient temperature prior to testing. The test specimen is placed between two plan parallel loading platens. The test specimen is then loaded by a rectangular and periodical vertical stress pulse, with a frequency of 0.5 Hz and a load of 100±2 kPa. The cumulative axial strain is calculated after 1000 loading cycles

^£ 1000 = 100-( ^M1 °.°°) , where Ewoo is the cumulative axial strain of the test specimen after 1OOO loading cycles in percent, u oo is the cumulative permanent deformation of the test specimen after 1000 cycles, in millimetres to the nearest 0.01 mm and tj is the initial thickness of the test specimen in millimetres.

The loading is stopped if the cumulative axial strain exceeds 4% and the specimen is considered demolished or failed and the number of cycles is reported as cycles to failure.

The results are seen from Table 14 for triplicate analysis for the reference sample and samples with digestate. Outlier removal was conducted for the reference and Sample 1.

Table 14: Cycles to failure and cumulative axial strain after 1000 cycles, Ewoo, from tri-axial rutting for asphalt samples; reference sample with limestone filler and sample 1 to 12 with digestate replacing 50% filler in triplicate for each sample

Figure 6 shows cycles to failure and rutting at 1000 cycles according to En 12697-25:2016 of reference asphalt samples compared with digestate samples having 50% filler replaced with digestate and 6.2 wt% 70/100 penetration grade bitumen. In Figure 6, a box and whiskers plot of the tri-axial rutting test results is seen. Most of the tested samples require fewer cycles to fail than the reference, but not significantly different from the reference or out of the industry specifications. The rutting experienced after 1000 cycles is higher for the samples compared to the reference, but not significantly different. This is a consequence of the softening effect that the dried solid digestate added as 50% filler replacement has on the bitumen and can be countered by adjusting the asphalt composition and/or the bitumen quality and/or amount. It can also be used as an advantage to achieve a softer asphalt.

Example 7 Test of filler performance of the digestate active filler composition

The stiffness and phase angles are tested in the samples obtained from Example 3, Table 7 and 8. A reference asphalt AC11 surf asphalt sample with 6.2% 70/100 bitumen and 12 samples where 50% filler material was replaced with digestate Sample 1 to 12. The bitumen was extracted according to EN 12697-1 :2020, soxhlet extractor method, before rheologic assessment.

7.1 Master curve construction

From recording of linear viscoelastic rheological properties from small strain oscillatory testing using a dynamic shear rheometer (DSR), master curves at a reference temperature of 20°C are constructed for mastics in their unaged and aged conditions. The effect on stiffness is seen from the Complex shear modulus (G*) and for the effect on phase angle (p). On the master curve for effect on stiffness values below the master curve means that the tested sample is less stiff (i.e., softer) than the reference. A comparison with the master curve will show whether the tested samples provide an equally balanced effect over the whole width of the spectrum of complex shear modulus G* measured in Pascal (Pa) vs. angular frequency measured in radian per second (co or rad/s). In relation to the effect on phase angle, values above the master curve means that the tested sample comprises more liquid than the reference and such samples are accordingly less elastic than the reference.

If the samples shown an equally balanced effect over the whole width of the spectrum, it must be assumed that the samples were not subjected to polymer modification.

Figure 7 shows the master curves for reference mastic and average for mastics with 50% filler replacement with dried, milled digestate in unaged and aged form with 70/100 penetration grade bitumen

A) Master curves for Shear stress modulus (G*) for extracted bitumen from fresh and aged reference and samples with digestate, B) Master curves for phase shift angle (p) for extracted bitumen from fresh and aged reference and samples with digestate, C) Master curves for complex shear modulus and phase shift angle for extracted bitumen from fresh and aged reference and average for samples with digestate combined with 95% confidence interval. In Figure 7 are the master curve recorded at 20°C seen. The spectral values are averaged, and the 95% confidence interval is calculated and shown with dotted lines (Figure 7C). The It is observed that the effect is equally affected over the whole width of the spectrum, and thus, no effect equal to polymeric modification is observed.

In Table 15 are the Complex shear modulus and Phase shift angles at 1.59 Hz or 10 radians per second corresponding to a load mimicking the load from a heavy truck driving with 80 km per hour according to The Shell Bitumen Handbook, edition 6, page 141.

Table 15: Complex shear modulus and phase shift angles from DSR master curves for asphalt samples; reference sample with limestone filler and sample 1 to 12 with digestate replacing 50% filler in triplicate for each sample

7.2 Effect on Stiffness

The curves from all fresh samples provide the same shape as the fresh reference sample, thus, the tested samples are equally balanced over the whole width of the spectrum. All samples show values below the reference sample. Thus, the tested samples are less stiff /softer than the reference sample.

For the aged samples, the same tendency is found. Thus, the curves from all aged samples provide the same shape as the aged reference sample. Although the values are lower (less stiff/softer) than the reference aged sample, the samples are equally balanced over the whole width of the spectrum. The change in complex shear modulus is on average lower for the samples containing solid digestate after aging compared to the reference showing anti-aging properties. On average the aging index (Alfaqawi et al. 2022) or the complex shear modulus ratio at 10 radian per second or 1.59 hertz is 17% lower, being 8.3 for the reference sample and 6.9±1.4 for the samples with 50% filler replaced by dried solid digestate. This is seen in Figure 8 which shows a box and whiskers plot for complex shear modulus ratio between unaged and aged bitumen from reference bitumen extracted from standard mastic and bitumen extracted from mastics with 50% filler replacement with dried, milled digestate in unaged and aged form with 70/100 penetration grade bitumen. 7.3 Effect on Phase Angle

The curves from all fresh samples provide the same shape as the fresh reference sample as seen from Figure 7B. Thus, the samples are equally balanced over the whole width of the spectrum. All samples show values above the reference sample. Thus, the tested samples are more viscous or softer even after aging than the reference sample.

Example 8 Calcium content in solid digestate

In the inorganic fraction found from Example 2, Table 5, a significant portion of the ash or inorganic content consists of calcium, most likely calcium salts and complexes containing calcium. To investigate the amount of calcium content in the ash fraction an analysis was performed by drying the ash from solid digestate samples to total dryness at 60°C and grinding the sample to pass a 1 mm screen. Then the samples were dissolved in an open vessel with concentrated hydrochloric and nitric acid (aqua-regia) using a temperature-controlled block. The elements dissolved in the acid are analyzed by inductively coupled plasma - optical emission spectroscopy (ICP-OES). The calcium content of some samples accounts for approximately one fifth of the inorganic content and calcium salts and/or complexed thus accounts for a substantial amount of the dry matter of the solid digestate samples. As the dry digestate and digestate ash suspended in water forms bubbles upon addition of hydrochloric acid, calcium carbonates are present. If all calcium is assumed to be present as calcium carbonate, the calcium carbonate would amount to approximate half of the inorganic fraction of solid digestate. In Table 16 are the calcium concentration on dry matter basis for the digestate samples 1 , 2, 3, 4, 5, 6, 7, 8 an 9, respectively, described in Example 1 seen. The ash content is also seen as well as a calculation of how much of the ash or dry matter calcium carbonate would constitute of the digestate samples if it is assumed that all detected calcium is calcium carbonate.

Table 16: Analysis of calcium content in solid digestate samples by ICP-OES and calculation of % of dry matter and ash if all calcium is assumed to be calcium carbonate

At the Northwich Renescience plant, the solid digestate was sampled routinously every workday. 50 g of sample is sampled and stored in a plastic container in a refrigerator at 4°C. Samples are compiled over approximately two weeks until the container is full and contains approximately 500 g of sample. The combined sample representing the average of two weeks production of solid digestate was analyzed for calcium content. The calcium content on average was 123±40 g per kg dry matter of the solid digestate for 38 routine samples taken in the period from January 2021 to December 2022. If all calcium is calcium carbonate, the calcium carbonate would account for 31 ±10 wt% of the dry solid digestate or 55±12 wt% of the inorganic, ashsso’c, fraction.

Example 9 Lignin characterization

Solid digestate from treatment of MSW contains both structural carbohydrates and lignin due to the nature of the substrate. Compositional analysis was performed of 126 subsamples of 600 kg MSW sampled approx, every second week through more than two years of operation of the Renescience Northwich plant and hand sorting into categories such as plastic types, glass, metal types and paper, cardboard, and kitchen putrescibles. It has been found that the MSW used to produce solid digestate contains 4 to 15 wt% paper, 3 to 12 wt% cardboard, 2 to 9 wt% fines and 1 to 9 wt% putrescibles, all of which mainly is lignocellulosic biomass. The substrate for the enzymatic and/or microbial process thus contains from 1/10 to 1/2 lignocellulosic biomass-like material.

In Example 2.1 Chemical composition analysis, the solid digestate is analysed with compositional analysis methods developed for determining amounts of carbohydrates and lignin in biomass samples. When analysing lignocellulosic biomass, the acid insoluble fraction can be assumed to be Klason-lignin as no other fractions present are acid insoluble. In solid digestate, other components from the substrate might be acid insoluble. To further characterise the solid digestate, the content of lignin originating from lignocellulosic biomass is sought quantified and characterized.

9.1 Lignin quantification by pyrolysis-GC-MS

The lignin content originating from cellulosic materials such as grasses, softwood or hardwood of three dried spot samples of solid digestate was analysed by pyrolysis gas chromatography with mass spectrometric detection (py-GC/MS) according to Erven et al., ACS Sustainable Chem. Eng. 2019, 7, 20070-20076. The lignin was not isolated prior to analysis in order to quantify the proportion of lignin in the acid resistant components described in Example 2.1 , Table 4, where it was found that the dried solid digestate contains from 20 to 23 wt% of the dry matter acid resistant components for digestate Sample 1 , 2 and 3, which would be considered Klason-lignin in a pure biomass sample. A sample size of 80-90 pg was used. Through pyrolysis in a Multi-shot pyrolyzer (Frontier Laboratories, New Ulrn, MN, USA) equipped with an AS-1020E Autoshot autosampler, the samples are decomposed in the absence of oxygen at 500°C for 1 minute with an interface temperature of 300°C. The pyrolyzer was coupled to a Trace 1300 GC equipped with a Restek DB-1701 fused-silica capillary column (30 m x 0.25 mm i.d. 0.25 pm film thickness, Thermo Scientific). The pyrolysis products were injected on the column via a split injection at 250°C with a split ration of 1 :133 using helium as a carrier gas with a constant flow of 1.5 ml/minute. The GC oven was programmed to maintain 70°C for 2 minutes followed by an increase in temperature from 70°C to 270°C at a rate of 5°C per minute. The temperature was then held at 270°C for 15 minutes. This procedure was selectively separating the products into monomeric building blocks of lignin. The GC was coupled to an Extractive Orbitrap Mass Spectrometer from Thermo Scientific, quantitatively detecting the lignin products. The detection was conducted at an electron ionization (El) at 70 eV and a source temperature of 250°C and a scan range of m/z 50-550 and a scan rate of 4.0 scans/sec. Compounds were identified by comparing retention time and mass spectrum with standards, the NIST library and published data, further a reference sample of 13C-labelled wheat straw was included in the data set for comparison.

In Table 17, the acid resistant compounds in solid digestate, from Table 4, found from the NREL protocol are compared with the amount of lignin found by py-GC/MS. It was found that not all acid resistant components were derived from lignin. The quantification of lignin by py-GC/MS is underestimated due to not including all constituents of lignin and that the pyrolysis process is known to experience negative interference when used on samples with a very high ash content such as digestate samples. The acid resistant components method of quantifying lignin in biomass samples developed by NREL, Sluiter et al. (NREL/TP 510 42618): Determination of Structural Carbohydrates and Lignin in Biomass is known to overestimate lignin in some sample matrices. Protein complexes, plastic components, and possible other constituents from MSW ending up in solid digestate is also acid resistant. Further, some inorganic components can interfere with the acid solubilisation, some oils and waxes might be present and cause positive interference, some uncertainty is expected from the protein estimation and the Sluiter method is not recommended on samples with a high ash content such as solid digestate due to neutralisation of the acid. Thus, the true lignin in solid digestate is more likely closer to 5 - 6 wt% compared to 8.4 to 11.7 wt% of the dry matter in solid digestate found in the present analysis. Table 17: Acid resistant components analysed According to Example 2.1 , Table 4 and py-GC/MS relative abundance of structural features within ¹² C/ ¹³ C solid digestate samples 1 , 2 and 3 on the basis of molar peak area

Further, the nature of the lignin is analysed and the results is seen in Table 18.

Table 18: Py-GC/MS relative abundance of structural features within ¹² C/ ¹³ C wheat straw (WS) and solid digestate samples 1 , 2 and 3 on the basis of molar peak area

Lignin aromatic units

Compared to lignocellulosic biomasses such as wheat straw, the lignin in solid digestate is guaiacyl rich, showing over 80% on molar basis being G-units. In fact, the S/G ratio in the digestate lignin fraction is very low compared to S/G ratio of native lignocellulose biomass which could be a result of both the mixed origin of lignocellulose in MSW and limited degradation of lignin occurring in the anaerobic digestion. From the data, it can be concluded that some degradation or condensation of the lignin has occurred, as the CA-0 and the methyl abundance is high.

Table 19: Py-GC/MS relative abundance of structural features within ¹² C/ ¹³ C wheat straw (WS) and solid digestate samples 1 , 2 and 3 on the basis of molar peak area separation of plastics and metals contains both native celluose structures (e.g wood, kitchen putrescibles etc) and processed lignocellulose (e.g., paper, cardboard etc). The cellulose is expected to have undergone enzymatic solubilisation to solubilise, hydrolyse and loosen the lignocellulosic structures into particles transferred with the liquid stream to the AD as well as a (some) microbial degradation before being processed to solid digestate. However, larger lignin structures could follow the fractions that are separated from the liquid fraction that is transferred to anaerobic digestion from which the digestate is produced. From mass balance of the plant, 4 to 17 wt% paper, containing approx. 1 wt% lignin, of dry matter according to Zhou et al. (2017), Lipids from waste paper, BioResources 12(3), 5249-5263 and 3 to 12 wt% carboard containing approximately 13 wt% lignin of the dry matter according to Zhou et al. (2017), Lipids from waste paper, BioResources 12(3), 5249-5263 would result in 4 to 16 wt% lignin of the solid digestate dry matter assuming that all lignin remains in the solid digestate and is not degraded. From the S/G ratio, it is observed that the lignin remaining in the digestate is not native lignin as for example wheat straw, but more processed lignin, most likely from paper and carboard. It is shown by Sammons et al 2013. Organosolv lignin analysis. Bioresources 8(2), 2752 - 2767, that lignin from newspaper has very low S/G ratio and it is known that cardboard is often made from softwood, which is largely composed of G units, supporting that the lignin in the digestate is mainly from the paper and cardboard in the MSW substrate rather than from native biomass such as wheat straw. The amount of paper and cardboard in the substrate would lead to approximately 17 wt% lignin on average, ranging from 7-30 wt%, of digestate dry matter if all paper and cardboard is transferred to the AD process, which it is not. This supports the assumption that approx. 5% of the digestate dry matter is lignin as found from the analysis in this Example.

9.2 Lignin characterisation

An alkaline extract was prepared by extracting 1 g of dry digestate in 8 mL 2M NaOH and heating the solution at 120 °C for 1 hour before quenching the reaction in cold water. The supernatant was isolated by centrifugation, acidified to pH 2 by adding 5M HCI to precipitate the lignin. The lignin was washed twice with 40 mL water and dried at 50°C.

The alkali-isolated lignin was characterized by HSQC NMR and high intensity signals was obtained, showing decent amounts of native lignin isolated. The extraction yield was 59-96 wt% of the quantified lignin amount by py-GC-MS and 14-25% of the acid resistant components. Further, very little variation between three monthly spot samples was observed showing consistent results regardless of the relative inhomogeneous substrate. The spectra is seen from Figure 10. Figure 10 shows the aromatic region (upper part of figure) and aliphatic region (lower part of figure) of HSEQ NMR spectra of alkaline lignin extracts from dried digestate Sample 1 , 2 and 3.

Example 10 Quantification of microplastics in digestate active filler composition

In Example 2.1 the digestate is analysed with compositional analysis methods developed for determining amounts of carbohydrates and lignin in biomass samples. When analysing lignocellulosic biomass, the acid insoluble fraction can be assumed to be Klason-lignin as no other significant fractions is present that is acid insoluble. In digestate, other components from the substrate might be acid insoluble, such as protein complexes and some plastic fragments. To further characterise the digestate, the content of lignin originating from lignocellulosic biomass in digestate is sought quantified and characterized, and the result is shown in Example 9, Table 17. It is expected that some plastic will go into the digestate fraction during processing. In this example, it is sought quantified how much. 10.1 Acid resistant components content in plastic

Plastic of various types are examples of fractions which may be acid insoluble. To illustrate that plastic is acid resistant, like Klason-lignin, two types of plastic was analysed according to the method described in Example 2.1 with no extraction. The samples were taken from food packaging going to household waste, consisting of coloured polypropylene (PP), and low-density polyethylene (LDPE). The samples were dried in an oven at 40°C over night, cut to smaller pieces and grounded on a Retsch mill with a 1 mm sieve. The analysis was performed in duplicate and the result is seen from Table 20.

Table 20: Anhydro carbohydrates, acid resistant components and ash content of plastic samples

10.2 Plastic quantified and identified through manual picking of particles larger than 1 mm

Impurities such as micro plastic and stones can be determined in sludge, treated biowaste and soil according to DS/CEN/TS 16202:2013. To investigate the amount of plastic in the digestate samples, this method was adapted for analysis of microplastics in digestate samples. Approximately 100 g of dry matter sample was weighed of into a 500 mL glass beaker, a subsample of 10 g was used for determining the dry matter content by drying overnight at 60°C in a convection oven from Binder, Germany, model FD53230V-9010-0082. 250 mL of tap water was added to the sample and the mixture was stirred regularly for at least one hour to allow agglomerates or other components to dissolve or soften, before adding a 20 mm magnetic stirring rod and stirring at 200 rpm until all visible lumps are dispersed. Sieves and lids (bottom and top) were dried over night at 60°C and weighed before assembling the sieve tower in the sieve shaker, a Vibratory Sieve Shaker AS 200 control 100- 240V, 50/60 Hz from Ninolab A/S. After assembling of the shake tower, the water hose was attached to the lid and a water faucet and tightened. A sample container was placed to collect the wash water and the water was turned on at a flow of 1 L/min. The sample was then added to the top sieve and was spread out with a glass rod, followed by shaking the sieve tower at 2 mm/g amplitude for 5 minutes. The shake tower was subsequently disassembled and if lumps were formed, they were dispersed with the glass rod. This was repeated 4 times in total. The shake tower was disassembled, each sievedried overnight at 60°C and weighed to quantify the mass in each sieve fraction. Three different digestate samples were analysed for particle size distribution by wet sieving as described above. The results are seen from Table 21.

Table 21 : Particle size distribution found from wet sieving of three digestate samples.

★calculated as the remaining dry matter not found on the sieves

It is observed that the fraction of particles smaller than 63 pm is slightly larger when wet sieving compared to analyzing the particle size distribution by laser diffraction on dried samples. This is expected as agglomeration during the drying process is avoided. Table 22: Comparison of fraction of particles passing 60-63 pm when analyzing un-milled wet sample or milled or un-milled dry samples

* See Table 1

Figure 11 shows pictures of larger wet and dry sieve fraction of digestate Sample 10. It is seen that plastic pieces are clearly visible in wet sieved samples. A: 1 mm sieve fraction after wet sieving of Sample 10, B: 1 mm sieve fraction after dry sieving of Sample 10, C: 2 mm sieve fraction after wet sieving of Sample 10, D: 2 mm sieve fraction after dry sieving of Sample 10 (no material in this fraction). The fractions are spread on a white piece of paper and sorted manually with a set of tweezers according to DS/CEN/TS 16202:2013 into glass, metals, plastics and other materials. The plastic is identified by IOSYS mlRoGun2.0 plastic identifier (IR tech) and sorted into PET, PS, PA, PVC, PP. Dark plastic is identified visually as IR technology cannot detect dark plastic types. Figure 12 shows A: unsorted material from dried 2 mm sieve fraction of digestate, Sample 4, and B: sorted and identified material from dried 2 mm sieve fraction of sample 4 showing plastic identified by IR, dark plastic and miscellaneous fibers and agglomerates. From Table 23, it is seen that of the larger particles, the 1 and 2 mm sieve fractions, plastic particles can visually be identified and manually sorted with tweezers to quantify that 22-90% based on weight of the larger particles consists of plastic particles

Table 23: Particle size distribution for 1 and 2 mm sieve fractions found from wet sieveing of solid digestate samples with amount of said fractions manually identified as plastic

PVC, PET, PP and PS was positively identified by IR in all samples analyzed. Dark plastics that cannot be identified by IR was also found in all samples, as was miscellaneous fibers and granules. Further other plastic types such as PLA, PEPA, PCA, PPPT, PO, ABS and OS with other additives was indicated in the samples.

10.3 Plastic analysis by LD-IR

Two samples, Sample 4 (Nov-21) and Sample 9 (Apr-22) were analysed for microplastics in particles with a size down to 20 pm. Subsamples of the two samples were air dried at 60°C before analysis to determine dry matter content. Approximately 2 grams of dried samples were oxidated with Fenton’s reagent, a mixture of hydrogen peroxide and ferrous iron, to remove organic material. The material is then density separated in with calcium chloride to remove the bulk inorganic mineral fraction. The extract from the density separation is passed through a 5 pm filter and the retained particulates are transferred onto an infra-red reflective slide and analysed in a two-step process.

First, the number of particles and particle size of the particles is identified by a direct infra-red imaging (LDIR) system for particles from 20 to 500 pm. An infrared spectrum of each particle is recorded in the plastic fingerprint region of 975-1800 cm ^-1 to provide a chemical identification based on comparison of each spectra with a database of reference spectra.

Particles larger than 500 pm and smaller than 5 mm is identified with Attenuated Total Reflection (ATR), where nine polymers: polyethylene (PE), polypropylene (PP), polystyrene (PS), polyvinylchloride (PVC), polyethylene terephthalate (PET), polycarbonate (PC), polymethylmethacrylate (PMMA), polyamide (PA) and polyurethane (Pll), can be identified routinously with certainty, where the IR spectral identification has been validated with external standards. Further, acrylonitrile butadiene styrene (ABS), polylactic acid (PLA) and Polytetraflouroethylene (PTFE) is identified as well as rubber particles, where the IR spectral identification has not been validated with external standards.

When analysing microplastics in waste waters, a certainty of 80% spectral match is used. In order to ensure a conservative estimate when identifying the amount of microplastics in digestate samples it was decided to apply a certainty of 70%. Even when using this conservative estimate, an explanation degree of more than 80% of all particles in the cleansed samples was achieved. The number of plastic particles in each size category is seen from Table 24.

Table 24: Micro plastic particles identified in Sample 4 and Sample 9 with validated plastic types in white and unvalidated plastic types in grey In Table 25 are the percentage of each identified plastic type seen. Table 25: Validated identified plast types PE, PP, PS, PVC, PET, PC, PMMA, PA and Pll, as well as not validated identified plast particles of ABS, PLA, PTFE and rubber in % of microplastic particles found in the two digestate samples.

Plastic types [% of particles found in 0.5 g sample)

The amount of microplastic can be estimated for, for example, Pll by assuming quadratic form of the particles with a hight, length and width as the larger of the particle size range. For example will 175 Pll particles of a size of 200-500 pm have an approximate diameter of 300 pm volume of 2.7-1 O’ ¹¹ m ³ ' 175 particles = 4.7-10’ ⁹ m ³ . Pll can have densities ranging from 0.3 to 1.2 kg/m ³ as it is used in a range of applications from coatings to foam applications. If the highest density of 1.2 is used to give the most conservative estimate of weight based Pll in the sample, the 2431 Pll particles in the sample is estimated to amount to 0.005 wt% of the dry matter for Sample 4 and the 697 particles in sample 9 amounts to 0.003 wt% of the dry matter.

In Table 26 are a summary of the known constituents of the acid resistant components. It is seen that 2-5% is not accounted for. Lignin from py-GC-MS might be slightly underestimated and will account for some. Plastic is not quantified in the particles smaller than 20 pm and between 500-1000 pm and will account for some. If the same plastic content as the 1-2 mm fraction is assumed for the 0.5-1 mm fraction, approx. 0.5 and 0.9 wt% plastic of dry matter is found here. This plastic can be the source of some of the ‘other’ fraction. The remaining is most likely caused by overestimation of acid resistant components due to a small interaction from the high inorganic content as previously discussed.

Table 26: Summary of details for acid resistant components

* Slightly underestimated due to not analysing all components of lignin

** Particles not cleansed and weight might be overestimated

*** Methods optimised towards waste water and soils with very small sample amount, which might exclude larger particles

Example 11 : Extraction of apolar components in dichlormethane and identification thereof

To further characterize the digestate, an extraction with dichlormethane (DCM), which is routinely used to solubilise and extract bitumen in asphalt mixtures, was carried out and the extract was characterised. For the characterization 660 g in total of an equal mixture of digestate Sample 1 , 2 and 3, described in Example 1 , was mixed with 3 L of DCM and left to extract under a N2 headspace at room temperature for 24 hours. The extraction was continued under reflux for another 4 hours in a Soxhlet extractor to further extract more material. Subsamples were collected for analysis before the solution of DCM with digestate after extraction were centrifuged, and the supernatant was left to dry at room temperature in a laboratory fume hood to quantify that approx. 2.6% of the digestate dry matter was extracted. The obtained DCM extract was characterised by 2D Heteronuclear Single Quantum Correlation (HSQC) NMR, High Performance Liquid Chromatography (HPLC) and Gas Chromatography with Flame Ionization Detection (GC-FID) with fatty acid methylesterifiation (FAME) derivatization and Gas chromatography coupled with Mass Spectrometry GC-MS without derivatisation.

Analysis with 2D HSQC NMR of the DCM extracts which were obtained at room temperature and reflux, showed that, in the samples extracted under reflux more compounds were extracted compared to the room temperature extract (Figure 13). Figure 13 shows the total HSQC NMR spectra of digestate DCM extracts (left: Ambient temperature extraction and right: Extraction under reflux) showing carbon stretch (SC) on the F1 axis and hydrogen-strech (5H) on F2 axis. However, neither of the DCM extracts showed presence of lignin-related substructures in the spectra. Strong signals in the aliphatic (CH2/CH3) region, which are associated with vinyl groups of fatty acids (5C/5H 129.3/5.3 ppm), were observed, suggesting that fatty acids and related compounds were abundantly present in both the DCM extracts obtained. For HPLC analysis, approx. 10 mg extract was mixed with 1 mL milli-Q water and 1 mL 1 M H2SO4 containing 100 mM valeric acid as internal standard. Samples were vortexed and mixed on a rotator at room temperature for 30 minutes, centrifuged and filtered through a 0.2 pM cellulose filter, before being injected on a Shodex Rspak KC-811 column. HPLC analysis of the sample extracts did not show the presence of volatile fatty acids, glycerol or monomeric saccharides.

For GC-FID, approx. 10 mg extract was weighed in a 5 mL conical glass tube, to which 2 mL 15% H2SO4 in methanol and 2 mL chloroform containing 0.5 mg/mL methyl pentadecanoate as internal standard were added. Samples were incubated for four hours under continuous stirring at 87°C, subsequently cooled on ice for five minutes and transferred to 15 mL Greiner tubes. 1 mL of milli-Q water was added, and samples were briefly vortexed and centrifuged. The bottom chloroform layer was transferred to GC-vials and analysed by using a Zebron ZB-FAME column. GC-FID after FAME derivatization confirmed the presence of various saturated and unsaturated long-chain fatty acids, in particular palmitic (C16:0), stearic (C18:0) and oleic (C18:1) acid (Figure 14, Table 27). Figure 14 shows GC-FID analysis of FAME derivatized DCM extracts of ambient temperature extract (top) and reflux extract (bottom). Quantification of these fatty acids showed a combined content of 40.6 and 65.4 mg/g of the extract at room temperature and under reflux, respectively, indeed highlighting that the extract obtained through reflux is slightly higher in fatty acids. Many other, long(er) chain fatty acids and other apolar compounds not matching the retention times of commercial standards available were present in the extract, together corresponding to the majority of the peaks and peak area present.

Table 27: GC-FID quantification of identified fatty acids

For GC-MS analysis, FAME derivatized samples prepared for GC-FID analysis were used and in addition an underivatized sample was analysed. For the underivatized sample, approx. 1 mg of the combined extract was dissolved in 1 mL DCM. Injected compounds were separated by using a Rxi- 5ms column with a Trace 1300 GC coupled to a Thermo Scientific single quadrupole ISQ-7000 MS. The GC-MS analysis of the FAME derivatized and underivatized DCM extract confirms the presence of a myriad of apolar components, as seen from the GC spectra in Figure 15. Figure 15 shows GC- MS analysis of derivatized DCM extract of ambient temperature extract (A) and reflux extract (B) and underivatized combined DCM extracts (C).

Example 12: Calculation of organic biogenic carbon content in dry digestate/active filler and carbon removal/CC footprint

Digestate active filler produced and described in Example 1 , contains biogenic carbon in range from 100 to 400 kg per ton of dry matter. The range can be even broader depending on the waste composition and the efficiency of the conversion of organic material in the AD process. To calculate the CO2 footprint, organic (biogenic) carbon was measured by the DUMAS method. The result is seen in Table 28 for the digestate samples used throughout the examples.

Table 28: Total organic carbon in solid digestate Sample 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 and 12 quantified by the DUMAS method

At the Northwich Renescience plant, the solid digestate was sampled routinely every workday. 50 g of sample was sampled and stored in a plastic container in a refrigerator at 4°C. Samples were compiled over approximately two weeks until the container was full and contains approximately 500 g of sample.

The combined sample representing the average of two weeks production of solid digestate was analyzed for organic carbon according to the DUMAS method described in Example 2. It was found that the organic carbon content on average was 22±5 wt% of the dry matter of the solid digestate for 37 routine samples taken in the period from January 2021 to December 2022. The variation observed in the data set was from 14 to 31 wt% organic carbon of the dry matter of solid digestate, as seen from Figure 9. Figure 9 shows the organic carbon and estimated protein content from DUMAS analysis of routine samples of digestate from the Renescience Northwich plant for a 2 year period where the date of sampling is the day of analyzing the compiled sample from two weeks of daily sampling.

According to standard: Sustainability of construction works - environmental declarations - Core rules for the product category of construction products, EN 15804, 1 kg biogenic carbon is equivalent to 44/12 kg of CO2 equivalent. The corresponding biogenic CO2 equivalent content in digestate active filler is between -100*44/12=-367 to -400*44/12= -1467 kg CO2 equivalent per ton digestate active filler, which is gross biogenic CO2 equivalent content.

To understand the environmental impact of digestate active filler and quantify the potential carbon removal, net biogenic CO2 equivalent content needs to be calculated by subtraction of CO2 equivalent emissions during the active filler production process.

A Life Cycle Assessment (LCA) study was performed, where the following emissions were identified: Emissions during transportation of waste Digestion process emissions Emissions due to enzyme consumption Emissions due to electricity consumption Emissions due to heat used in digestion Emissions due to polymer use Emissions because of heat used in drying process Emissions due to transportation to asphalt producer

The above emissions from the enzymatic and/or microbial process followed by AD that can be allocated by mass to digestate active filler production from the process including energy recovery from biogas used in a CH P (combined heat and power) unit are approximately 60 kg CO2 equivalent per ton digestate active filler. The emissions may increase in case fossil fuel sources for energy production are used or decreased in case enzyme consumption or transportation emissions are reduced.

Net biogenic CO2 equivalent removed can be calculated by subtracting the emissions during digestate active filler production activities from the biogenic CO2 equivalent content in digestate active filler. This provides a net negative CO2 emission in range from -300 to -1400 kg CO2 equivalent per ton digestate active filler.

Net biogenic CO2 equivalent calculated above can potentially be a basis for entering CO2 removal market.