TECHNICAL FIELD

The present invention relates to a method and a device for heating of a liquid food product.

BACKGROUND

In modern liquid processing systems heat treatment is often desirable for making the final product stable during subsequent processing and storage. Such heat treatment, i.e. pasteurization, is normally performed by heating the liquid product to an elevated temperature, which temperature is sufficient to at least prevent microbial growth in the liquid product, as well as keeping the liquid product at that particular temperature for a specific period of time before the liquid product is cooled.

There is an established practice among fruit juice manufacturers and bottlers that liquid product such as juices are commonly pasteurized twice before reaching the consumer. As mentioned above the purpose of these heat treatments is to make the juice product stable during its planned storage period. A first pasteurization is done as soon as possible after juice extraction, or as a first step in the evaporator. An example of the primary pasteurization is at 95 °C for 30 sec. The second pasteurization is carried out prior to filling the juice in its container. A conventional example of the second pasteurization is to keep the liquid product at 95 'Ό for 15-30 sec.

SUMMARY

It is, therefore, an object of the present disclosure to provide a method and a device for heat treating a liquid food product.

To this end an object is to provide a method for heat treating a liquid food product comprising directing the liquid food product into a heat exchanger, leading the food product through the heat exchanger, heating and maintaining the liquid food product at a temperature of about 80 °C for < 15 s, and/or obtaining a pasteurization unit (PU) of at least 0.05 min as determined according to Formula (I):

PU = t x 10 ^(" ^ ^} (I), wherein t is the holding time in minutes, T is the effective holding time, °C, z is the temperature, in °C, and obtaining a pasteurized liquid food product.

In some aspects an object is to provide a treatment about 80 °C for 15 s. In some aspects an object is to provide the method further comprises directing the liquid food product into the heat exchanger at a first product temperature (t _M ), and allowed to exit at a second product temperature (t ₀ i ), and the method further comprising directing a heating medium into the heat exchanger at a first heating medium temperature (t _i2 ), leading the heating medium through the heat exchanger and allowing it to exit the heat exchanger at a second heating medium temperature (t _o2 ), wherein the temperature difference (dT) between the liquid food product and the heating medium exceeds 5 °C.

Additionally an object is to provide a control unit configured to perform the method for heat treating a liquid food product comprising

directing the liquid food product into a heat exchanger, leading the food product through the heat exchanger, heating and maintaining the liquid food product at a temperature of about 80 °C for < 15 s, and/or obtaining a pasteurization unit (PU) of at least 0.05 min as determined according to Formula (I), wherein t is the holding time in minutes, T is the effective holding time, °C, z is the temperature, in °C, and obtaining a pasteurized liquid food product.

Additionally an object is to provide a heat exchanger comprising the control unit configured to perform the method for heat treating a liquid food product comprising directing the liquid food product into a heat exchanger, leading the food product through the heat exchanger, heating and maintaining the liquid food product at a temperature of about 80 °C for < 15 s, and/or obtaining a pasteurization unit (PU) of at least 0.05 min as determined according to Formula (l),wherein t is the holding time in minutes, T is the effective holding time, °C, z is the temperature, in °C, and obtaining a pasteurized liquid food product.

BRIEF DESCRIPTION OF DRAWINGS

The above, as well as additional objects, features, and advantages of the present invention, will be better understood through the following illustrative and non- limiting detailed description of preferred embodiments of the present invention, with reference to the appended drawings, wherein:

Fig. 1 is a schematic view of a pasteurization system;

Fig. 2 is a schematic view of a heat exchanger in which embodiments may be applied, as well as illustrative temperature profiles; and

Fig. 3 is a view similar to that of Fig. 3 of yet a further embodiment.

DETAILED DESCRIPTION

In an effort to put the present disclosure into its proper context reference is first made to Figs. 1 and 2.

Starting with Fig. 1 a prior art pasteurization system 10 is shown. The system

10 includes a batch tank 1 1 enclosing a certain amount of liquid product to be heated. Upon heating, the liquid product is transported through a heat exchanger 12 wherein the temperature of the liquid product is elevated to a predetermined pasteurization temperature. The liquid product is thereafter kept at the elevated temperature during transportation through a holding cell 13 for ensuring the desired pasteurization. After pasteurization is completed, the liquid product is typically transported through a further heat exchanger 14 for cooling down the liquid product. During heating, a temperature sensor 15 provides a measurement signal representing the current temperature of the liquid product. Additionally, a flow meter 16 provides a measurement signal representing the current flow of the liquid product. The measured temperature is compared with a reference value in a control unit 17 for determining if the actual liquid product temperature is within an allowed pasteurization temperature interval.

Correspondingly, the measured flow is compared with a reference value in a control unit 18 for determining if the actual liquid product flow is within an allowed

pasteurization temperature interval.

As described herein there is a need for heat treatment of juices, nectars and still drinks, which is essential for obtaining the desired product shelf life. The process should additionally be designed to minimise unwanted degradation of constituents in the product in order to maintain the desired quality and to minimise changes in quality as the product may be stored long periods of time, for example a year or more. Loss of quality can lead to reduced product quality, foaming during deaeration and filling, and uneven distribution of floating pulp in packages. Chapter 4 of The Orange book, printed 2004, ISBN 91 -3428-4 discloses many considerations of concern to those skilled in the art of treating and packaging juices, nectars and still drinks.

Orange juice, as well as most other juices, is pasteurised at least twice before it reaches the consumer (except for a small amount of NFC being filled directly into consumer packages). The first pasteurisation occurs immediately after extraction prior to bulk storage, and the second pasteurisation occurs before packaging. The first pasteurisation is necessary to inactivate the enzyme pectin methyl esterase (PME) which may otherwise cause cloud loss in liquid food products such as juice, nectars and still drinks. This is of even more concern when the liquid food product is a concentrate. Microorganisms of commercial interest, e.g. yeast, pathogens, mould etc are also killed by this process. The second pasteurisation destroys any

microorganisms that may have (re-)contaminated the liquid food product after the first pasteurisation step and survived bulk storage, as well as those which may have (re- )contaminated the liquid food product during its reconstitution from concentrate. As one example the concentrate is diluted with potable water.

It can be concluded that the pasteurization process is essential to obtain commercially useful liquid food product having a long shelf-life, and it is of most importance to perform each step in a way to secure that none of the issues discussed herein above will take place before consumption of the liquid food product. It is also of concern not to damage the liquid food product in a way that compromise the quality of the product, for example the consumer will most likely react negative to a change in the liquid food product, for example change in colour or taste of an pre-existing liquid food product. The risks are often associated with inadequate pasteurization.

In some aspects the pasteurization is to inactivate enzymes from the fruits and to inactivate microorganisms in the liquid food product. For example in the case of orange juice the first pasteurisation is necessary to inactivate the enzyme pectin methyl esterase (PME), which will cause cloud loss of single strength juice and gelation of concentrate during storage.

Some aspects relates to a method for producing a liquid food product having been sufficiently treated, e.g. pasteurized. In some aspects the method relates to improvements compared to conventional processes as according to aspects disclosed herein it allows less process changes, for example when changing liquid food product processed by a system suitable for performing the methods according to one or more aspects and embodiments disclosed herein. In some aspects the methods lead to lower energy consumption. The lower energy consumption is generally of much interest in view of increasing energy costs and environmental concern. In some aspects the methods lead to improved liquid food products as the heat load applied to the liquid food product can be optimized and in many embodiments improved compared to conventional processes. In some aspect the methods enable less process variations.

Calculations using the herein described methods can generate a 19% energy cost reduction per year. The calculation was based on a reference line processing 22,000 litres of orange juice per hour in a Tetra Therm Aseptic Drink (16 hours/day, 5 days/week, 50 weeks/year, 1 hour cleaning/day). This additionally results in a 20% improvement in carbon footprint.

In some aspects the methods relate to providing a medium to be pasteurized, heat treating the medium to a first temperature, maintaining the first temperature, heat treating the medium to a second temperature, and maintaining the second temperature and obtaining a liquid food product.

Some aspect described herein relates to the second heat treatment being at a temperature about 80 'Ό for <15 s.

Some aspect described herein relates to the second heat treatment being at a temperature about 80 °C for 15 s. Some aspect described herein relates to the second heat treatment being at a temperature 80 °C for 15 s.

The first pasteurization is not essential for certain products such as certain still drinks where the second heat treatment is sufficient to provide the desired sterilization.

Generally the first pasteurization step is performed close to the harvesting site, or even at the harvesting cite, i.e. juice extract is collected can be divided into juice from concentrate (FC) , juice not from concentrate (NFC), and freshly squeezed orange juice. Freshly squeezed orange juice is not pasteurized and thus have a rather short period in which it should be consumed. FC and NFC are generally subjected to a first pasteurization to inactivate/kill pectin methyl esterase, as well as other organisms. The juices are after the pasteurization optionally concentrated in order to obtain FC. FC is for example stored aseptically or frozen, for example at temperatures from -6 to - 25 °C. The demands depend on the type and quality of the juice and are within the capacity of the skilled person to control. NFC is after the pasteurization stored frozen, for example in drums at -18 °C or lower; or chilled (in large containers at around -1 to 1 °C).

Generally any pasteurization temperature and time conventionally used for the particular liquid food product, i.e. necessary to achieve the desired level of

pasteurization, depend on the quality of the fruit and fruit extract. For example orange juices are pasteurized between 95-98 °C for 10-30 s. Generally the pasteurization should not be less than 72°C for 15 s as otherwise pathogenic bacteria such as Salmonella, Listeria or E.coli may be present.

After storage and optional transport the FC or NFC is heated (reconstituted FC) and thereafter subjected to pasteurization. The pasteurization may be the second pasteurization the liquid food product is subjected to. The main purpose of a second pasteurization is microbial destruction, which for example could be from the thawing or from the reconstitution process as well as from transportation. In some embodiments the pasteurization is performed at 80 'C for 15s, or at a corresponding heat treatment such as 78°C for 30 s. A method for pasteurization of a liquid food product thus comprises directing the liquid food product into a heat exchanger, leading the food product through the heat exchanger, heating and maintaining the liquid food product at a temperature of about 80 °C for < 15 s, and/or obtaining a pasteurization unit (PU) of at least 0.05 min as determined according to Formula (I):

PU = t x 10 ^(" ^ ^} (I), wherein t is the holding time in minutes, T is the effective holding time, °C, z is the temperature, in °C, and obtaining a pasteurized liquid food product.

The low temperature and short time have surprisingly been found to be sufficient for many liquid food products, such as juice, nectar and still drinks. For some drinks the first pasteurization is not necessary, for example for still drinks. In some embodiments the method of pasteurization of liquid food products, such as juice, nectars and still drinks disclosed herein are having a pH of equal to or less than 4.2, for example less than 3.8.

In some embodiments the method relates to pasteurizing liquid food products such as juice, e.g. FC or NFC; nectars; and still drink all having a pH < 4.2. pH values disclosed herein are generally determined by pH meter having an accuracy of +/- 0.05

In some embodiments the method relates to pasteurizing juice, such as FC or NFC; nectars; still drink having a pH < 4.2 at 80 'C for < 15s, such as 15 s. Generally The temperatures disclosed herein are generally determined by temperature sensors having a precision of < +/- 0.25 °C.

Accordingly some aspects relate to obtaining a pasteurized liquid food product having a pH < 4.2, which pH is maintained during the pasteurization, according to the methods described above. The obtained pasteurized liquid food product is thus possible to obtain at a desired or even improved quality as the low temperature can lead to less degradation, colour changes and taste changes all depending on the composition of the liquid food product.

In some embodiment the liquid food product is a liquid comprising organic acids. In some embodiments the liquid food product is a liquid comprising one or more organic acid selected from the group consisting of lactic, citric and/or malic acid. In some embodiments the liquid food product is a liquid comprising one or more organic acid selected from the group consisting of citric and/or malic acid. In some

embodiments the liquid food product is a liquid comprising organic acid selected from the group consisting of citric and/or malic acid and has a pH < 4.2.

In some embodiments of the present invention the liquid food product comprises at least 3.5 g/L of organic acids. In some embodiments of the present invention the liquid food product comprises at least 1 .5 g/L malic acid, such as at least 2, 4, 6, and 8 g/L malic acid, and/or at least 1 .5 g/L citric acid, such as 2, 4, 6, and 8 g/L citric acid. In some embodiments of the present invention the liquid food product comprises at least 1 .5 g/L lactic acid, such as at least 2, 4, 6, and 8 g/L lactic acid, The organic acid content can be determined in accordance with standards described in CODEX STAN 247-2005. The citric acid content and malic acid content is determined in accordance with CEN 1 137-1994 and CEN 1 138-1994 respectively. The lactic acid content can be determined in accordance with CEN 12631 -1999.

In some embodiments as described herein the liquid food product should not contain spores of Alicyclobacillus in 10 g / 10 ml (i.e. there should be no detectable spores in 10 g or 10 ml of the liquid product).

In some embodiments as described herein the liquid food product should not contain ascospores of mould such as Byssochlamys in 10 g / 10 ml.

The presence of Alicyclobacillus can be determined in accordance with IFUMB12 - 2007 and Byssochlamys can be in accordance with IFUMB04 - 1996

In some embodiments a turbulent flow is an advantage.

In some aspects the method for pasteurization of a liquid food product can be performed in a system as described in Fig.2 which illustrates a heat exchanger 12 which may be used to realize the present invention according to one or more embodiments thereof. Fig. 2 is schematic and it will also be used to describe a general operation of a heat exchanger as well as some key parameters in such operation. Apart from the constructional illustration in the lower portion of Fig. 2, the upper portion of Fig. 2 illustrates the temperature profiles for the product as well as for the heating medium as it passes through the heat exchanger, also in a schematic manner.

In Fig. 2 the product to be heated enters from the right and flows to the left through the central pipe 27, whereas the heating medium enters from the left and flows to the right in the surrounding pipe 28. The illustrated heat exchanger 10 thus utilizes a countercurrent flow, which is particularly beneficial for the purposes as described herein. Apart from the constructional illustration in the lower portion of Fig. 2, the upper portion of Fig. 2 illustrates the temperature profiles for the product as well as for the heating medium as it passes through the heat exchanger, also in a schematic manner.

A shorts summary of the denotation used in Fig. 2 reads:

t _M - product temperature when entering heat-exchanger step.

t ₀ i - product temperature when leaving heat-exchanger step.

t _i2 - temperature of heating medium when entering heat-exchanger step. t ₀ 2 - temperature of heating medium when leaving heat exchanger step

At _m - temperature difference between heating medium and product as a function of position.

The product entering from the right will have a lower temperature t _M

(compared to t ₀ i ) and as it passes through the inner pipe it will gradually absorb heat from the heating medium conducted through the walls of the inner pipe, and consequently the heating medium will be cooled as it travels from the left to the right. By balancing the flows it is possible to maintain an almost constant temperature difference At _m (dT in the following for increased readability) between the product and the heating medium for each position in the heat exchanger. A reason for obtaining an as constant dT as possible may be that it may be desired to have a maximum dT while not exceeding a maximum value. A constant dT will then make it possible to obtain an optimal effect. Depending on the flows dT will be higher in the region of the product inlet or in the region of product outlet. For a co-flow heat exchanger, which may be exemplified by switching the flow direction of one of the flows in Fig. 2, it is apparent that the maximum dT is found at the product inlet of the heat exchanger, where the temperature of the product is as low as possible and the temperature of the heating medium is as high as possible.

In one or several embodiments the heating medium may consist of a product to be cooled, thus forming a regenerative heat exchanger with even more beneficial energy efficiency, this may be referred to as product/product heating (or cooling, since the product being the heating medium will be cooled correspondingly, see in particular the upper portion of Fig. 2). In alternative related embodiments the temperature of the heating medium may be adjusted up or down before entering the heat exchanger.

Given that Fig. 2 is schematic it should be emphasized that inventive features may be applied to heat exchangers of various designs. In an actual design the heating may be conducted in several steps, and the product/product heating (or cooling) may e.g. be used in a first heating step or an intermediate heating step, wherein the last heating is performed using another heating media, such as hot water or steam to boost the temperature up to a desired temperature. The preferred relationship for dT may be applicable for any heat exchanger in this steps. A counterflow heat exchanger may preferred for maintaining a constant temperature difference dT, however within that specification there are numerous types of heat exchangers to chose from, such as plate heat exchangers, tube heat exchangers or scraped surface heat exchangers. A typical tube heat exchanger would, in contrast to the one of Fig. 2, typically comprise several parallel pipes for leading the product, even though Fig. 2 only illustrate a single pipe in each direction. A typical tube heat exchanger installation is shown in

WO0031489 to the present applicant. Further, a co-flow heat exchanger could also be utilized.

In the embodiment illustrated in Fig. 3 further features have been added. In this embodiment the product, once having reached the desired temperature, is guided to a holding cell. In the holding cell the product is maintained at a specific temperature or within a specific temperature interval for a specific period of time. The holding cell may be a container or vessel, yet for a well defined process it may be preferred for the holding cell comprises a specified length of pipe. It should be noted that there may be additional heat exchangers or heaters arranged to boost the temperature of the product before it enters the holding cell. In the holding cell a predetermined

temperature t _H c is maintained for a predetermined period of time T _H c- According to one aspect t _H c is defined by75 °C < t _H c < 95 °C and wherein the predetermined period of time T _HC is about 20 s or less. According to one specific embodiment t _H c is about 80 °C and T _HC is <15.

Most heat exchanger systems used in liquid food processing are designed and dimensioned for processing of dairy products, and the same systems are utilized for juice, nectars, still drinks etc. For this reason the systems of today operates at a temperature difference of about 3-5 'Ό. In the present invention a method for heating a liquid product has been optimized for juice, nectars and still drinks having a pH of about 4.2 or less, for example 3.8 or less. Moreover the process is according to one or several embodiments limited to a second pasteurization, i.e. the pasteurization performed immediately before filling into consumer containers (as opposed to the first pasteurization which is generally performed prior to storage of the product).

Benefits of the method according to one or more of the embodiments disclosed herein are that the increase temperature difference dT results in a more efficient heat transfer process, which reduces the energy consumption. The surprising result that the product quality in accordance with the used indicators was not affected by the quite large variation in dT (compared to the conventional 3-5 °C) as compared to a baseline also results in an increase versatility. Various products within the field of juices, nectars and still drinks may have different properties in terms of viscosity, specific heat etc, all affecting a heat transfer process. Instead of tailor making systems for each of these products use may be made of a variation of dT. This means that transitions between operating conditions when changing from a first product to a second may be made swiftly and with ease.

In some embodiments the pasteurization as described herein, is performed in a counterflow heat exchanger, and the liquid food product is directed into the heat exchanger at a first product temperature (t _M ), and allowed to exit at a second product temperature (t ₀ i ), which temperatures (t _M , t ₀ i ) may be the same or different. The method further comprises directing a heating medium into the heat exchanger at a first heating medium temperature (t _i2 ), leading the heating medium through the heat exchanger and allowing it to exit the heat exchanger at a second heating medium temperature (t _o2 ) which temperatures (t _i2 , t _o2 ) may be the same or different, wherein the temperature difference (dT) between the liquid food product and the heating medium exceeds 5 °C. In some embodiments the method further comprises redirecting the obtained pasteurized liquid food product through the heat exchanger, now as the heating medium, other examples of heating medium are water and steam.

In some embodiments the dT is > X≥ Y °C, wherein X independently of Y is selected from the group consisting of 5, 6, 7, 8,9, 10, 1 1 , 12, 13, 14, 15, 16, 17,18, 19, 20, 21 , 22, 23, 24, 25, 26, 27, 28, 29, 30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44, 45, 46, 47, 48, 49, and Y independently of X is selected from the group consisting of 6, 7, 8,9, 10, 1 1 , 12, 13, 14, 15, 16, 17,18, 19, 20, 21 , 22, 23, 24, 25, 26, 27, 28, 29, 30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44, 45, 46, 47, 48, 49, 50.

As used herein the term "juice" is naturally contained in fruit or vegetables. It is prepared by mechanically squeezing or macerating fresh fruits or vegetables. Juice is 100 % fruit juice. Examples of juices are orange and apple juice, another example is juices containing juice from multiple fruits, for example orange/pineapple.

As used herein the term "nectar" refers to fruit or vegetables but with a 25-99 % juice content and usually with added sugar. Examples of nectar are orange nectar, and nectar containing multiple fruits such as apple, pineapple, orange, and

passionfruit.

As used herein "still drinks" contain 0-24 % juice content in fruit, vegetable or other flavours. Example of still drinks are energy drinks such as Powerade marketed by the Coca Cola Company, ice tea, fruit drinks such as Festis marketed by Carlsberg.

As used herein "D-value" refers to decimal reduction time and is the time required at a specified temperature to kill 90% of the organisms being studied. When referring to D-values it is proper to give the temperature as a subscript of the "D".

As used herein "z-value" is a term used in thermal death time calculations. The z-value of an organism is the temperature, in degrees Celsius, that is required for the thermal destruction curve to move one log cycle. It may be simplified as the temperature required for one log reduction in the D-value. While the D-value gives the time needed at a certain temperature to kill an organism, the z-value relates the resistance of an organism to differing temperatures.

As used herein in connection with specified temperatures "about" is intended to mean +/- 3 °C. Generally whenever a specific temperature or interval or

temperatures are given without the wording "about" there is a tolerance allowing the temperature to vary slightly such as +/- 1 °C.

As used herein a "heat exchanger" is a system built for energy transfer from one medium to another. A number of experiments described in detail below where performed in order to verify the invention and secure that the proposed pasteurization method described herein works. Several species known to have been involved in spoilage of foods were used, see for example Gaze, J.E. 2006. Pasteurisation: a food industry practical guideline. Guideline No. 51 (Second edition). Campden & Chorleywood Food

Research Association Group, UK. ISBN-10:0-905942-89-2. ISBN-13:978-0-905942-89- 6.

The species selected were Bacillus megaterium, Bacillus licheniformis, Bacillus coagulans, Paenibacillus macerans, Paenibacillus polymyxa, Clostridium butyricum and Clostridium pasteurianum.

Examples

Bacterial strains

Bacillus megaterium A 4764-25D; Bacillus licheniformis M 3644-3; Bacillus coagulans DSM 2356 and Paenibacillus macerans M 2455-3C_SPC were obtained from a culture collection at Tetra Pak® in Stuttgart, Germany. Paenibacillus polymyxa CCUG 7426 (type strain); Clostridium butyricum CCUG 4217 (type strain) and Clostridium pasteurianum CCUG 31328 (type strain) were obtained from Culture Collection University of Goteborg, Sweden. The bacteria were kept on tryptone soya agar (TSA, Merck) plates. All strains were streaked on orange serum agar (OSA, Oxoid) with a pH of 5.5, to check the potential to grow in the fruit juices. This medium is used for production control in the fruit juice industry.

Preparation of spore suspensions

Spore suspensions were prepared on different kinds of agar plates. Bacillus licheniformis was streaked on nutrient agar plates (Difco) supplemented with 0.01 M MnCI ₂ (manganese (II) chloride); 0.20 M MgCI ₂ (magnesium (II) chloride); 0.14 M CaCI ₂ and incubated at 30 ^< C for 1 1 days. Almost 100% of the cells sporulated.

Paenibacillus polymyxa was streaked on orange serum agar (OSA) plates and incubated at 30 °C for nine days. About 90% of the cells formed spores. Clostridium butyricum was streaked on reinforced clostridial medium (RCM) agar plates (Oxoid) and incubated anaerobically at 30 °C for 14 days. About 80% of the cells formed spores, but very few of the spores were released from the mother cells. Clostridium pasteurianum was streaked on RCM-agar plates and incubated anaerobically at 30 °C for 14 days. About 80% of the cells formed spores, but very few spores were released from the mother cells. Anaerobic incubation was made in anaerobic jars with

Anaerocult (Merck).

The spores were harvested by pouring ice cold sterile distilled water on the plates and rubbing the spores of the surface with a T-shaped spreader. The spores were transferred to a test tube and washed three times with ice cold sterile distilled water by centrifugation at 5000 rpm for 15 minutes. The spore suspensions were heat treated to inactivate vegetative cells (80 ^/10 min B. licheniformis, CI. butyricum, CI. pasteurianum; ySOSmin P. polymyxa). The number of spores was determined and the suspensions were kept at refrigeration temperature until further diluted in sterile deionised water and used for inoculation of juice packages.

Growth of vegetative bacteria

B. megaterium, B. coagulans and P. macerans failed to form spores and were grown stationary as vegetative bacteria in Tryptone Soya Broth (Oxoid) to a cell density of about 10 ⁷ cfu/ml. The cells were diluted in sterile 0.9% NaCI before adding to the juice packages.

The tests were performed in commercial ready to drink 250 ml portion packs of orange and apple juice at different pH by aseptically inoculating the bacterial suspension into 10 packages of apple juice and 10 packages of orange juice. Prior to the inoculation juice was withdrawn aseptically through the existing straw hole of each package and the pH adjusted using sterile sodium hydroxide or hydrochloric acid. The hole was covered with paraffin to get it tight again. The packages were shaken to distribute the added base or acid and the bacteria or spores evenly in the packages before incubation. Bacteria or spores were added in 0.1 ml portions to give a load of about 100 cfu/ml in the packages. Five replicates were done for each bacterial species, pH and juice product, respectively. In total 280 samples were inoculated, incubated and analysed for growth and pH. The inoculated packages were incubated at room temperature (20-23 for three weeks with some exceptions. Two packages from each pH-group and juice product inoculated with CI. butyricum and CI. pasteurianum, respectively, were incubated anaerobically. Two packages from each pH-group and juice product inoculated with B. coagulans were incubated at 37 ^< C for another week after the three weeks incubation at room temperature, before opening.

Analysis of growth

The packages were opened aseptically and a 10 μΙ sample was withdrawn with a loop and streaked on the surface of an agar plate. B. licheniformis and P. polymyxa were streaked on both PCA and OSA and incubated at 30 °C for 5 days. CI. butyricum and CI. pasteurianum were streaked on RCM-agar and incubated anaerobically at 37 ^< C for 5 days. B. megaterium, P. macerans and B. coagulans were streaked on TSA. B.

megaterium and P. macerans plates were incubated at 30 'C for 5 days and B.

coagulans plates were incubated at 37 ^< C for 5 days.

The pH in each package was determined and the content of the package was poured into a glass beaker to inspect possible growth of moulds caused by

recontamination at pH adjustment or inoculation of bacteria or spores.

After heat treatment at 80 °C for 10 min the number of B. licheniformis and P.polymyxa spores were determined on 3M Petrifilm for total count to be 5.2x10 ⁹ cfu/ml and >10 ⁸ cfu/ml, respectively. The number of CI. butyricum and CI. pasteurianum spores were determined on RCM-agar, anaerobically, to be 7.7x10 ⁶ cfu/ml and 3.0x10 ⁶ cfu/ml, respectively.

The pH of the samples were about pH 3.5, 3.8, 4.0 and 4.2.

B. licheniformis, P. polymyxa, CI. butyricum and CI. pasteurianum were inoculated into the fruit juices as spores. None of them grew in any of the juices in any of the pH-groups. The colony morphology was studied and the bacterial cells were examined in microscope. They corresponded well to the species inoculated. Thus the spores can be recovered from the juice in the examined pH range, but they cannot germinate and form growing vegetative cells. No other growth was detected in any of the packages.

B. megaterium, B. coagulans and P. macerans failed to form spores on any of the tested media and were inoculated into the fruit juices as vegetative cells. None of them grew in any of the juices in any of the pH-groups. The colony morphology was studied and the bacterial cells were examined in microscope. They corresponded well to the species inoculated. The vegetative bacterial cells could be recovered from the juice in the examined pH range, but they could not continue to grow. No other growth was detected in any of the packages, showing that no contamination was made during pH adjustment and inoculation of spores.

The finding in the example above concluded that none of the above sporeforming bacteria grew in pH < 4.2, which would thus mean a potential to reduce the heat treatment currently used, i.e. 95-98 "C for 10-30 s, as the heat treatment doesn't have to consider sporeforming bacteria such as those above. In order to verify the finding above and make commercial use thereof, theoretical calculations of a target of 9 log reduction of yeast ascospores, as well as large scale tests were completed.

The theoretical calculations where based on apple juice ¹ as prepared according to the description below having a D ₆₃ -value of 1 .6 min and z-value of 5.4 ^< Ό, a buffert solution ² at a pH 4.5 having a D ₆ o-value of 22 min and z-value of 6.5 °C, and orange juice ³ at a pH 3.8 having a D ₆ 5-value of 22 min and z-value of 5.5 'Ό. Table 1 shows the result of the calculation and the pasteurization temperature. The buffert solution referred to was used as it illustrates one known scenario.

The test was designed to find what treatment temperature and holding time is necessary in order to produce a microbiologically shelf-stable liquid food product (i.e. 3 weeks at 20-23 °C) taking into account resilient spores such as the ascospores. In order to fulfil criteria of providing a shelf-stable liquid food product yeast ascospores were added to apple juice made from frozen concentrate diluted with water to °Brix of 12, the juice having a pH of 3.5. 2x10 ⁷ ascospores per 300 litre apple juice (pH 3.5) were added. The juice was pasteurized at three different temperatures and maintained at this temperature for 15 s. The juice was aseptically packaged in 250 ml Tetra Brik® Aseptic carton packages, which were stored at room temperature (20-23 'Ό) for 3 weeks and inspected for swollen or blown packages which was taken as a general indication of sufficient pasteurization was achieved. Swollen packages were opened aseptically and detection of the test organism performed by general means.

Table 1 . Results of tests and calculations. Based on the above it was concluded that the treatment at 80 'Ό and holding time of 15 s did provide the desired level of treatment, and sufficient number of log reductions, 14, even on the most heat resistant ascospores.

In order to make further verification a batch of 4,000 litres of orange juice (1 1 .3 ^Q Brix, pH 4.0) was pasteurized at 78°C/22 s, which corresponds to a heat treatment at 80 5 s, i.e. a lower heat treatment than 80°C/15 s. The juice was packed in 16,000 Tetra Prisma ® Aseptic packages (250 ml). The packages were stored at room temperature (20-23 °C) for 3 weeks prior to inspection. None of the packages had any sign of gas formation due to unsterility. 1 ,043 packages were transported to an internal laboratory at Tetra Pak in Lund where they were opened and streaked at orange serum agar. All of them were commercially sterile. The result confirms that 80 ^Q C for <15 s may give enough heat load in the second pasteurization to produce a commercially sterile product.