GLUCOSE LEVELS

FIELD OF THE INVENTION

[0001] The present invention relates to improvement of symptoms of Type 1 Diabetes using compositions derived from Phyllanthus emblica, or extracts thereof. In an embodiment, post-prandial blood glucose levels may be reduced in a patient afflicted with Type 1 Diabetes. This invention also relates to a method of improving endothelial function and cardiovascular health using such compositions.

BACKGROUND

[0002] Many herbs possess potent antioxidant, anti-inflammatory and cardio-protective properties and are used by patients with increased risk of cardiovascular morbidity and mortality in order to treat or prevent disease and/or reduce symptoms. Among them, Phyllanthus emblica, syn. Emblica officinalis Gaertn., the Indian gooseberry (PE, "Amla") is widely used in Indian medicine for the treatment of various diseases. There are studies which show significant anti-hyperglycaemic and lipid lowering effects of PE in diabetic patients. In in-vitro and animal studies, PE demonstrates potent antioxidant effects against several test systems such as superoxide radical and hydroxyl radical scavenging action, and in systemic augmentation of antioxidant enzymes in animals (Antony, et al, "A pilot clinical study to evaluate the effect of Emblica officinalis extract (Amlamax™) on markers of systemic inflammation and dyslipidemia," Indian J. Clin. Biochemistry (2008) 23(4): 378-381).

[0003] PE has traditionally been used in rasayana formulations, a group of plant-derived drugs that are reputed to promote health and longevity by augmenting the body's defense mechanisms against disease, arresting the aging process, revitalizing the body in debilitated conditions, increasing the capability of the individual to resist adverse environmental factors and creating a sense of mental well-being.

[0004] Capros® is a standardized aqueous extract of the edible fruits of Phyllanthus emblica (Emblica officinalis, Indian Gooseberry or "Amla"). It has been shown in human clinical studies in type 2 diabetic subjects as well as subjects with metabolic syndrome that Capros® decreases glycosylated hemoglobin (HbAlc) levels significantly, indicating that it helps treat type 2 diabetes (P. Usharani, et al, Diabetes, Metabolic Syndrome and Obesity: Targets and Therapy (2013) 6:275-284).

[0005] There are other studies on certain Amla extracts which show similar activity. The study by Akhtar, et al., International J. Food Sciences and Nutrition (2011) 62(8): 609-616, evaluated the anti-hyperglycemic and lipid-lowering properties of Emblica officinalis Gaertn. fruit in normal and diabetic human volunteers. The results indicated a significant decrease (P < 0.05) in fasting and 2 hour post-prandial blood glucose levels on the 21st day in both normal and type 2 diabetic subjects receiving 1, 2 or 3 g E. officinalis powder per day as compared with their baseline values. Significant (P < 0.05) decreases were also observed in total cholesterol and triglycerides in both normal and diabetic volunteers on day 21 that were given either 2 or 3 g E. officinalis powder per day. However, diabetic volunteers receiving only 3 g E. officinalis powder exhibited a significant (P < 0.05) decrease in total lipids on day 21. Both normal and diabetic volunteers receiving 2 or 3 g E. officinalis powder significantly (P < 0.05) improved high-density lipoprotein-cholesterol (HDL-C) and lowered low-density lipoprotein-cholesterol (LDL-C) levels.

[0006] The antioxidant properties of amla extracts and their effects on the oxidative stress in streptozotocin-induced diabetes were examined in rats (Rao, et al, J. Medicinal Food (2005) 8(3): 362-368). Amla in the form of either the commercial enzymatic extract SunAmla (Taiyo Kagaku Co. Ltd., Yokkaichi, Japan) (20 or 40 mg/kg of body weight/day) or a polyphenol-rich fraction of ethyl acetate extract (10 or 20 mg/kg of body weight/day) was given orally for 20 days to the streptozotocin-induced diabetic rats. Amla extracts showed strong free radical scavenging activity. Amla also showed strong inhibition of the production of advanced glycosylated end products. The oral administration of amla extracts to the diabetic rats slightly improved body weight gain and also significantly alleviated various oxidative stress indices of the serum of the diabetic rats. The elevated serum levels of 5- hydroxymethylfurfural, which is a measure of glycosylated protein that is an indicator of oxidative stress, were significantly reduced dose-dependently in the amla-fed diabetic rats. Similarly, the serum level of creatinine, yet another oxidative stress parameter, was also reduced. Furthermore, thiobarbituric acid-reactive substances levels were significantly reduced with amla, indicating a reduction in lipid peroxidation. In addition, the decreased albumin levels in the diabetic rats were significantly improved with amla. Amla also significantly improved the serum adiponectin levels. These results thus form the scientific basis supporting the efficacy of amla for relieving the oxidative stress and improving glucose metabolism in diabetes.

[0007] Although there is enough data in the literature to show that amla extract helps reduce blood glucose levels in type 2 diabetics, it is not known to reduce post-prandial blood glucose levels in type 1 diabetic subjects. If a way could be found to reduce post-prandial blood glucose levels in human subjects (adult or juvenile) afflicted with type 1 diabetes mellitus, this would represent a contribution to the medical arts. [0008] Further, if a way could be found to use a therapeutically effective amount of Phyllanthus emblica, or extract thereof, in a composition to treat or prevent endothelial dysfunction and/or diabetes, this would represent a useful contribution to the medical arts.

SUMMARY

[0009] In one embodiment, the present invention describes the usefulness of Phyllanthus emblica and compositions or extracts thereof, in improving symptoms and/or metabolic outcomes in patients with Type 1 diabetes mellitus as well as in healthy subjects.

[0010] A method of treating or preventing Type 1 diabetes mellitus (T1DM) is provided including administering to an individual in need thereof a therapeutically effective amount of a composition comprising an extract of Phyllanthus emblica, wherein a blood glucose level is improved in that a lower post-prandial blood glucose level is obtained. The improvement may include lowering of post-prandial blood glucose to a range of about 70 mg/dl to about 200 mg/dl.

[0011] Other embodiments are contemplated for the effective treatment of human patients having Type 1 diabetes mellitus (T1DM). In one embodiment, a method of lowering postprandial blood glucose in an individual afflicted with Type 1 diabetes mellitus includes administering to an individual in need thereof a therapeutically effective amount of an extract of Phyllanthus emblica wherein a post-prandial blood glucose level is improved. The improvement may include lowering of post-prandial blood glucose to a range of about 70 mg/dl to about 200 mg/dl.

BRIEF DESCRIPTION OF THE DRAWINGS

[0012] FIG. 1A illustrates post-prandial blood glucose levels in a juvenile patient having Type 1 diabetes mellitus in a baseline experiment (each line representing a separate daily determination) wherein a shaded horizontal bar parallel with the x-axis (time) represents a preferred blood glucose range in mg/dl (about 70-150 mg/dl).

[0013] FIG. IB illustrates post-prandial blood glucose levels in a juvenile patient having Type 1 diabetes mellitus in one embodiment in accordance with the present invention wherein the patient was treated with Capros® (250 mg) orally, 10 minutes before a breakfast meal on three different days. As in FIG. 1 A the shaded horizontal bar parallel with the x-axis (time) represents a preferred blood glucose range in mg/dl (about 70-150 mg/dl).

[0014] FIG. 2 illustrates post-prandial blood glucose levels in a juvenile patient having Type 1 diabetes mellitus in one embodiment in accordance with the present invention wherein the patient was treated with Capros® (250 mg) orally, 10 minutes before a breakfast meal on at least four different days. As in FIG. 1 A the shaded horizontal bar parallel with the x-axis (time) represents a preferred blood glucose range in mg/dl. Y-axis represents blood glucose in mg/dl (about 70-150 mg/dl).

DETAILED DESCRIPTION

[0015] The following references are hereby incorporated by reference in their entirety, U.S. Patent Nos. 6,124,268, 6,290,996, 6,362,167, and 8,962,576. These references describe the isolation or preparation of extracts and extract blends as used herein.

[0016] In one aspect, the present invention reveals the usefulness of Phyllanthus emblica and compositions or extracts thereof, in improving symptoms and/or metabolic outcomes in patients with Type 1 diabetes mellitus as well as in healthy subjects.

[0017] As used herein, the term "prandial" means of or relating to a meal, i.e. ingestion or consumption of food by a human subject. In this context, "preprandial" or "pre-prandial" means before a meal, while "postprandial" or "post-prandial" means after a meal.

[0018] One suitable composition used herein is an extract blend which is isolated in stable form from the fruit of the Phyllanthus emblica plant, as described in detail in U.S. Pat. 6,124,268. The extraction process includes treating the finely-pulped fruit with a dilute aqueous or alcoholic-water salt solution, e.g. a 0.1 to 5% (w/w) sodium chloride solution, or the like, preferably at about 70 °C ± 5°C, or with a buffer solution, e.g. 0.1 to 5% (w/w) of sodium citrate/citric acid, or the like, filtering and drying, to provide the extract in powder form.

[0019] The extract includes the active constituents Emblicanin-A and Emblicanin-B, which are gallic/ellagic acid derivatives of 2-ketoglucono-5-lactone, in an amount by weight of about 35-55%, as well as: Punicagluconic acid, also named punigluconin (about 4-15% by weight), Pedunculagin (about 10-20%) by weight), Rutin (about 5-15% by weight), and low- to-medium molecular weight tannoids of gallic/ellagic acid (about 10-30%) by weight), gallic acid (up to about 5% by weight), and ellagic acid (up to about 5% by weight). Note that taken together, the overall extracted mixture or blend comprises an isolatable, identifiable, and purifiable group of components comprising a group of low molecular weight hydrolyzable tannoids ("LMwHTs"), generally excluding free gallic acid and ellagic acid. The composition may further include a nutraceutically or pharmaceutically acceptable carrier.

[0020] In one suitable embodiment, the amount of LMwHTs contained in a purified and/or enriched extract of Phyllanthus emblica is at least about 60% by weight. In another embodiment, the amount of LMwHTs contained in a purified and/or enriched extract of Phyllanthus emblica is greater than about 60% by weight. In another embodiment, the amount of LMwHTs contained in a purified and/or enriched extract of Phyllanthus emblica is greater than about 70% by weight. CAPROS® (available from Natreon, Inc., New Brunswick, New Jersey) is one such exemplary extract of Phyllanthus emblica.

[0021] The Phyllanthus emblica extract composition can be administered in a daily dosage from about 100 mg per day to about 3000 mg per day. In one preferred embodiment, the daily dosage is about 250 mg. In another suitable embodiment, the daily dosage is about 500 mg.

[0022] Patients with diabetes have vascular complications and endothelial dysfunction is one of the early prognostic markers of atherosclerosis which may eventually result in cardiovascular disease. Studies have reported that endothelial dysfunction occurs in patients with diabetes much earlier than clinical manifestations of diabetic vascular complications (Schalkwijk, et al., "Vascular complications in diabetes mellitus: the role of endothelial dysfunction," Clinical Science (2005) 109: 143-159). Diabetes is associated with accelerated atherosclerosis and microvascular complications are a major cause of morbidity and mortality, although in Type 1 , it is not clear whether endothelial dysfunction is a feature of the diabetic state itself. Endothelial cell dysfunction is emerging as a key component in the pathophysiology of cardiovascular abnormalities associated with diabetes mellitus.

[0023] The methods described above may be further understood in connection with the following Examples. The results of an extraction process may depend upon the solvent used, temperature of extraction, pressure at which extraction is performed, and duration of the extraction process. In several embodiments of this invention, these factors can be optimized to isolate and/or enrich and/or preserve the bioactives of Phyllanthus emblica.

[0024] According to the American Diabetes Association (2014) http://www.diabetes.org/living-with-diabetes/treatment-and-c are/blood-glucose- control/checking-vour-blood-glucose.html, blood glucose target for diabetic individuals are individualized. However, the American Diabetes Association suggests the following targets for most nonpregnant adults with diabetes. More or less stringent goals may be appropriate for each individual. Furthermore, the Association now recommends that children under the age of 19 diagnosed with type 1 diabetes strive to maintain an AIC level lower than 7.5 percent. http://www.diabetes.org/newsroorn/press-releases/2014/diabet es-association-sets- new-alc-target-for-children-with-type-l-diabetes.html.

[0025] AIC: 7%. AIC (7%) may also be reported as eAG: 154 mg/dl.

[0026] Before a meal (preprandial plasma glucose): 70-130 mg/dl. [0027] 1-2 hours after beginning of the meal (Postprandial plasma glucose): less than 180 mg/dl. Postprandial glucose may be more targeted if AIC goals are not met despite reaching preprandial glucose goals.

EXAMPLE 1 (Juvenile subject)

[0028] The present disclosure in one embodiment describes a case study in which Capros®, a standardized extract of Phyllanthus emblica, reduced post-prandial blood glucose levels in an 8 year old type 1 diabetic female patient.

[0029] The subject in this case study is an 8 year old girl who is a type 1 diabetic. Her care includes use of a continuous glucose monitor (CGM). When the CGM detects higher than set glucose level, she is given insulin injection to bring the glucose levels within normal limits.

[0030] One exemplary target blood glucose range is from about 75 to about 150 mg/dl in blood. This preferred range can be viewed as a shaded horizontal bar within the Figures.

[0031] In this case the breakfast meal was taken at about 8:00 AM. As shown in Fig. 1 A, post-prandial blood glucose levels increased significantly with this patient on several separate days after monitoring over several hours. This served as a baseline monitoring result.

[0032] The patient was administered one capsule of Capros® (250 mg) orally, 10 minutes before a breakfast meal on three different days. In contrast to the initial baseline result, the post-prandial blood glucose levels were lowered significantly, moderating within or generally toward the target blood glucose range over the course of about 3 hours (Fig. IB). In this case the breakfast meal was taken at about 8:00 AM.

EXAMPLE 2 (Juvenile subject)

[0033] The same patient of Example 1 was administered one capsule of Capros® (250 mg) orally, 10 minutes before a breakfast meal on at least four different days. In contrast to the initial result, the post-prandial blood glucose levels were lowered significantly, moderating within or generally toward the target blood glucose range over the course of about 4-5 hours (Fig. 2).

[0034] Examples 1 and 2 clearly demonstrated the improvement in post-prandial blood glucose levels after administration of Capros®. Based on the results, it is believed that Capros®, a standardized extract of Phyllanthus emblica fruit, reduced post-prandial blood glucose levels in the patient afflicted with type 1 diabetes.

[0035] Human clinical studies can be carried out in a randomized, double-blind, parallel and placebo-controlled study. Such clinical studies are being planned to confirm the findings described in Examples 1 and 2. It is expected that treatment of type 1 diabetic patients (juvenile or adult) with extracts derived from PE, such as Capros® (250 mg or 500 mg) will produce results as in Examples 1 or 2. That is, oral administration of Capros® before a meal is expected to provide post-prandial blood glucose lowering. One preferred blood glucose range is from about 70 to about 200 mg/dl. . Another suitable blood glucose range is from about 70 to about 150 mg/dl. Another suitable blood glucose range is from about 75 to about 130 mg/dl. It is further expected that AIC levels after administration of Capros® will be measured to be about 7.5% or less, or 7% or less.

[0036] It is also possible that Capros® may prove to reduce pre-prandial blood glucose levels also in type 1 diabetics, entirely replacing the need for insulin.

[0037] The nutraceutical compositions of the present invention may be administered in combination with a nutraceutically acceptable carrier. The active ingredients in such formulations may comprise from 1% by weight to 99% by weight, or alternatively, 0.1% by weight to 99.9% by weight. "Nutraceutically acceptable carrier" means any carrier, diluent or excipient that is compatible with the other ingredients of the formulation and not deleterious to the user. In accordance with one embodiment, suitable nutraceutically acceptable carriers can include ethanol, aqueous ethanol mixtures, water, fruit and/or vegetable juices, and combinations thereof.

[0038] The pharmaceutical compositions of the present invention may be administered in combination with a pharmaceutically acceptable carrier. The active ingredients in such formulations may comprise from 1% by weight to 99% by weight, or alternatively, 0.1% by weight to 99.9% by weight. "Pharmaceutically acceptable carrier" means any carrier, diluent or excipient that is compatible with the other ingredients of the formulation and not deleterious to the user.

[0039] Delivery system

[0040] Suitable dosage forms include tablets, capsules, solutions, suspensions, powders, gums, and confectionaries. Sublingual delivery systems include, but are not limited to, dissolvable tabs under and on the tongue, liquid drops, and beverages. Edible films, hydrophilic polymers, oral dissolvable films or oral dissolvable strips can be used. Other useful delivery systems comprise oral or nasal sprays or inhalers, and the like.

[0041] For oral administration, a Phyllantus emblica extract may be further combined with one or more solid inactive ingredients for the preparation of tablets, capsules, pills, powders, granules or other suitable dosage forms. For example, the active agent may be combined with at least one excipient such as fillers, binders, humectants, disintegrating agents, solution retarders, absorption accelerators, wetting agents, absorbents, or lubricating agents. Other useful excipients include magnesium stearate, calcium stearate, mannitol, xylitol, sweeteners, starch, carboxymethylcellulose, microcrystalline cellulose, silica, gelatin, silicon dioxide, and the like.

[0042] The components of the invention, together with a conventional adjuvant, carrier, or diluent, may thus be placed into the form of pharmaceutical compositions and unit dosages thereof. Such forms include solids, and in particular tablets, filled capsules, powder and pellet forms, and liquids, in particular aqueous or non-aqueous solutions, suspensions, emulsions, elixirs, and capsules filled with the same, all for oral use, suppositories for rectal administration, and sterile injectable solutions for parenteral use. Such pharmaceutical compositions and unit dosage forms thereof many comprise conventional ingredients in conventional proportions, with or without additional active compounds or principles, and such unit dosage forms may contain any suitable effective amount of the active ingredient commensurate with the intended daily dosage range to be employed.

[0043] The components of the present invention can be administered in a wide variety of oral and parenteral dosage forms. It will be obvious to those skilled in the art that the following dosage forms may comprise, as the active component, either a chemical compound of the invention or a pharmaceutically acceptable salt of a chemical compound of the invention.

[0044] For preparing pharmaceutical compositions from a chemical compound of the present invention, pharmaceutically acceptable carriers can be either solid or liquid. Solid form preparations include powders, tablets, pills, capsules, cachets, suppositories, and dispersible granules. A solid carrier can be one or more substances which may also act as diluents, flavoring agents, solubilizers, lubricants, suspending agents, binders, preservatives, tablet disintegrating agents, or an encapsulating material.

[0045] In powders, the carrier is a finely divided solid, which is in a mixture with the finely divided active component. In tablets, the active component is mixed with the carrier having the necessary binding capacity in suitable proportions and compacted in the shape and size desired.

[0046] The powders and tablets preferably contain from five or ten to about seventy percent of the active compound(s). Suitable carriers are magnesium carbonate, magnesium state, talc, sugar, lactose, pectin, dextrin, starch, gelatin, tragacanth, methylcellulose, sodium carboxymethlycellulose, a low melting wax, cocoa butter, and the like. The term "preparation" is intended to include the formulation of the active compound with encapsulating material as carrier providing a capsule in which the active component, with or without carriers, is surrounded by a carrier, which is thus in association with it. Similarly, cachets and lozenges are included. Tablets, powders, capsules, pills, cachets, and lozenges are included. Tablets, powders, capsules, pills, cachets, and lozenges can be used as solid forms suitable for oral administration.

[0047] Liquid preparations include solutions, suspensions, and emulsions, for example, water or water-propylene glycol solutions. For example, parenteral injection liquid preparations can be formulated as solutions in aqueous polyethylene glycol solution. The chemical compound according to the present invention may thus be formulated for parenteral administration (e.g. by injection, for example bolus injection or continuous infusion) and may be presented in unit dose for in ampoules, pre-filled syringes, small volume infusion or in multi-dose containers with an added preservative. The compositions may take such forms as suspensions, solutions, or emulsions in oily or aqueous vehicles, and may contain formulation agents such as suspending, stabilising and/or dispersing agents. Alternatively, the active ingredient may be in powder form, obtained by aseptic isolation of sterile solid or by lyophilization from solution, for constitution with a suitable vehicle, e.g. sterile, pyrogen- free water, before use.

[0048] Aqueous solutions suitable for oral use can be prepared by dissolving the active component in water and adding suitable colorants, flavors, stabilizing and thickening agents, as desired. Aqueous suspensions suitable for oral use can be made by dispersing the finely divided active component in water with viscous material, such as natural or synthetic gums, resins, methylcellulose, sodium carboxymethylcellulose, or other well known suspending agents.

[0049] Compositions suitable for topical administration in the mouth includes lozenges comprising the active agent in a flavored base, usually sucrose and acacia or tragacanth; pastilles comprising the active ingredient in an inert base such as gelatin and glycerine or sucrose and acacia; and mouthwashes comprising the active ingredient in suitable liquid carrier.

[0050] Solutions or suspensions are applied directly to the nasal cavity by conventional means, for example with a dropper, pipette or spray. The compositions may be provided in single or multi-dose form. In compositions intended for administration to the respiratory tract, including intranasal compositions, the compound will generally have a small particle size for example of the order of 5 microns or less. Such a particle size may be obtained by means known in the art, for example by micronization. [0051] The pharmaceutical preparations are preferably in unit dosage forms. In such form, the preparation is subdivided into unit doses containing appropriate quantities of the active component. The unit dosage form can be a packaged preparation, the package containing discrete quantities of preparation, such as packaged tablets, capsules, and powders in vials or ampoules. Also, the unit dosage form can be a capsule, tablet, cachet, or lozenges itself, or it can be the appropriate number of any of these in packaged form.

[0052] Tablets, capsules and lozenges for oral administration and liquids for oral use are preferred compositions. Solutions or suspensions for application to the nasal cavity or to the respiratory tract are preferred compositions. Transdermal patches for topical administration to the epidermis are preferred.

[0053] Further details on techniques for formulation and administration may be found in the latest edition of Remington's Pharmaceutical Sciences (Mack Publishing Co., Easton, PA).

[0054] Solid nutritional compositions for oral administration may optionally contain, in addition to the above enumerated nutritional composition ingredients or compounds: carrier materials such as corn starch, gelatin, acacia, microcrystalline cellulose, kaolin, dicalcium phosphate, calcium carbonate, sodium chloride, alginic acid, and the like; disintegrators including, microcrystalline cellulose, alginic acid, and the like; binders including acacia, methylcellulose, sodium carboxymethylcellulose, polyvinylpyrrolidone, hydroxypropyl methylcellulose, ethyl cellulose, and the like; and lubricants such as magnesium stearate, stearic acid, silicone fluid, talc, waxes, oils, colloidal silica, and the like. The usefulness of such excipients is well known in the art.

[0055] In one preferred embodiment, the nutritional composition may be in the form of a liquid. In accordance with this embodiment, a method of making a liquid composition is provided.

[0056] Liquid nutritional compositions for oral administration in connection with a method for preventing and/or treating inflammation, colds and/or flu can be prepared in water or other aqueous vehicles. In addition to the above enumerated ingredients or compounds, liquid nutritional compositions can include suspending agents such as, for example, methylcellulose, alginates, tragacanth, pectin, kelgin, carrageenan, acacia, polyvinylpyrrolidone, polyvinyl alcohol, and the like. The liquid nutritional compositions can be in the form of a solution, emulsion, syrup, gel, or elixir including or containing, together with the above enumerated ingredients or compounds, wetting agents, sweeteners, and coloring and flavoring agents. Various liquid and powder nutritional compositions can be prepared by conventional methods. Various ready-to-drink formulations (RTD's) are contemplated.

[0057] Routes of Administration

[0058] The compositions may be administered by any suitable route, including but not limited to oral, sublingual, buccal, ocular, pulmonary, rectal, and parenteral administration, or as an oral or nasal spray (e.g. inhalation of nebulized vapors, droplets, or solid particles). Parenteral administration includes, for example, intravenous, intramuscular, intraarterial, intraperitoneal, intranasal, intravaginal, intravesical (e.g., to the bladder), intradermal, transdermal, topical, or subcutaneous administration. Also contemplated within the scope of the invention is the instillation of a pharmaceutical composition in the body of the patient in a controlled formulation, with systemic or local release of the drug to occur at a later time. For example, the drug may be localized in a depot for controlled release to the circulation, or for release to a local site.

[0059] Pharmaceutical compositions of the invention may be those suitable for oral, rectal, bronchial, nasal, pulmonal, topical (including buccal and sub-lingual), transdermal, vaginal or parenteral (including cutaneous, subcutaneous, intramuscular, intraperitoneal, intravenous, intraarterial, intracerebal, intraocular injection or infusion) administration, or those in a form suitable for administration by inhalation or insufflations, including powders and liquid aerosol administration, or by sustained release systems. Suitable examples of sustained release systems include semipermeable matrices of solid hydrophobic polymers containing the compound of the invention, which matrices may be in form of shaped artices, e.g. films or microcapsules.

[0060] The use of the terms "a," "an," "the," and similar referents in the context of describing the presently claimed invention (especially in the context of the claims) are to be construed to cover both the singular and the plural, unless otherwise indicated herein or clearly contradicted by context. Recitation of ranges of values herein are merely intended to serve as a shorthand method of referring individually to each separate value falling within the range, unless otherwise indicated herein, and each separate value is incorporated into the specification as if it were individually recited herein. Use of the term "about" is intended to describe values either above or below the stated value in a range of approx. ±10%; in other embodiments the values may range in value either above or below the stated value in a range of approx. ±5%; in other embodiments the values may range in value either above or below the stated value in a range of approx. ±2%; in other embodiments the values may range in value either above or below the stated value in a range of approx. ±1%. The preceding ranges are intended to be made clear by context, and no further limitation is implied. All methods described herein can be performed in any suitable order unless otherwise indicated herein or otherwise clearly contradicted by context. The use of any and all examples, or exemplary language (e.g., "such as") provided herein, is intended merely to better illuminate the invention and does not pose a limitation on the scope of the invention unless otherwise claimed. No language in the specification should be construed as indicating any non-claimed element as essential to the practice of the invention.

[0061] While in the foregoing specification this invention has been described in relation to certain embodiments thereof, and many details have been put forth for the purpose of illustration, it will be apparent to those skilled in the art that the invention is susceptible to additional embodiments and that certain of the details described herein can be varied considerably without departing from the basic principles of the invention.

[0062] All references cited herein are incorporated by reference in their entirety. The present invention may be embodied in other specific forms without departing from the spirit or essential attributes thereof and, accordingly, reference should be made to the appended claims, rather than to the foregoing specification, as indicating the scope of the invention.