ENZYME PREPARATION FOR ACCELERATING THE AGING PROCESS OF CHEESE

Abstract: Disclosed is a composition and method for accelerating the aging process of cheese which involves the use of preserved, partially disrupted Lactobacillus casei or L. lactis, a similarly prepared concentrate of Lactobacillus plantarum and a blend of dried pregastic lipases.

Description: ENZYME PREPARATION FOR ACCELERATING TilE AGING PROCESS OF CHEESE BACKGROUND OF THE INVENTION This invention involves a composition and a method for the accelerated aging of American cheese, e.g. cheddar cheese, colby cheese, monterey and jack cheese. State of the art technology for accelerated cheese aging typically employs 1 of 2 approaches other than just raising the curing temperature. The first approach utilizes selected microorganisms in addition to the conventional cheesemaking ingredients whereas the second approach advocates the use of enzymes, in some cases from the same class of organisms, which are added during the cheese manufacturing process. Cheese flavor is a complex mixture of several hundred flavor components. The balance is especially critical to obtaining the typical organoleptic preception associated with cheese flavor. Also, associated with aging is body breakdown or softening. The complexity of the flavor and the opportunity for offflavor development have precluded the commercialization of an effective cheese aging product. Associated with the aging and flavor development of cheese is the overall hydrolysis of the major components thereof, i.e. the conversion of protein and fat to peptides and free fatty acids, respectively. Much focus has been placed on the role of proteolytic and lipolytic enzymes, e.g. lipases and esterases, from dairy microorganisms and from rennet during the ripening process. The formation of volatile fatty acids is routinely observed during the ripening process, however, the sources of these lipases and esterases are relatively undefined. Some authors assign lipolytic action to the milk lipase while others believe that contaminating microorganisms such as Micrococci sp. are the source of fat hydrolyzing enzymes. Current theories suggest that the flavor components of cheese are not necessarily the direct result of proteolysis and liopolysis, but instead are the result of precursor compounds generated by the gross hydrolysis of casein and fat which have been further converted by relatively obscure pathways. The balance of flavor components during cheese aging is a very delicate and complex process. Although the process is not well-understood, certain investigators have developed theories regarding the natural process and have consequently proposed methods to accelerate this process. Kratochvil discloses in U.S. patent 4,119,732 a process for the manufacture of American cheese which involves the addition of specific pairs of strains of XactobaciZZus plantarum and Streptococcus durans in combination with pre-gastric lipases during the cheesemaking process with subsequent curing of the cheese at elevated temperatures. In U.S. patent 4,158,607, Kalinowski, et al disclosed an enzymatic preparation composed of the protease from PeniciZZium candidum or P. roqueforti and the autolysate of a lactic acid bacteria from the species Streptococcus lactis or Lactobacillus casei. International Patent publication WO 82/03971 having an international publication date of November 25, 1982, discloses a method of producing a low-fat cheese product which involves inoculating milk with a culture of L. butgaricus and S. thermo phlus together with a culture of L. casei and a normal cheese starter culture. The first two organisms are said to produce the required flavor and rapid fermentation of lactose whereas the L. casei assists in the hydrolysis of protein. SUMMARY OF TIIE INVENTION The present invention is a composition for accelerating the aging process of cheese which comprises: a) a preserved, partially disrupted preparation of the lactic acid bacteria Lactobacillus casei, L. Zactis or a mixture thereof; b) a preserved culture concentrate of partially disrupted cells of Lactobacillus plantarum; and c) a blend of dried pre-gastric lipases. Also included within the scope of this invention is a method of accelerating the aging of cheese and/ or preventing the development of defects during the aging process using this preparation. DESCRIPTION OF THE INVENTION The cheese aging preparation of the present invention can be prepared by providing the three major ingredients in a premixed form so that they can be added to the milk during the cheesemaking process. Conversely, the ingredients may be added individually in whatever order is most convenient to the cheesemaker. The amounts and activity of the ingredients to be used are as follows: Ingredient Range % (W/W) Total Milk L. casei Preferred 0.025-0.03% at a standard* or protease level (140 PU/g) L. Zactis Operable 0.001-0.05% at a standard protease level (140 PU/g) *Standard Protease Unit (PU) = the amount of enzyme re quired to cause an in crease in adsorbance at 660 nm which is equivalent to the release of 0.1 pmole of tyrosine/hr/ml using a casein solution and the Folin reaction. Based on cell population before disruption L. plantarum Preferred 5 x 106 CFU/ml of milk** Operable 1 x 104 - 5 x 10 CFU/ml of milk **CFU = Colony Forming Units using ATP Agar (Difco Laboratories) for 48 hours at 37"C Grams/454 kg of milk Kid 4 Calf Preferred 0.30-0.35 g/454 kg of milk Lipases*** (kid) 0.13-0.18 g/454 kg of milk (calf) Operable 0.015-1.0 g/454 kg of milk (kid) Ingredient Range Grams/454 kg of milk (cont.) 0.0075-0.5 g/454 kg of milk (calf) ***Kid @ 2,500 2,500 RU/g Calf &commat; ; 5,500 RU/g Neutral Preferred 40,000-160,000 PU/454 kg Microbial of milk Protease Operable 0-600,000 PU/454 kg of milk RU = Ramsey units as determined by the assay described in Journal of Dairy Service, Vol. 50 (7) p. 1061 (1967). A selected strain of Lactobacillus casei or L. Zactis (preferably ATCC 39539 or ATCC 39538, respectively) is cultured in a hydrolyzed milk-based growth medium which is fortified with yeast extract and mineral salts. Cell biomass is produced during incubation for 16 to 24 hours at 37"C while the pH of the medium is controlled to a set point of 6.5 with NH40H additions. Cells are separated using centrifugation and the culture concentrate is subjected to mild disruption by several passes in a 2-stage homogenizer at 4,000 PSI or by other methods such as sonication autolysis or enzymatic-lysis such as by lysozyme addition. In a preferred embodiment, the preparation contains L. casei or L. Zactis cells of which approximately 67% are disrupted when subjected to disruption by standard techniques. Total cell disruption is not unsatisfactory but typically, the amount of disruption will be from 20% to 80% for both the L. casei, L. Zactis and L. plantarum described below. Any gentle method of preserving the partially disrupted product, such as freezing, freeze-drying or fluidized bed drying, is acceptable. Preferably, a suitable carrier is added and the complex is spray-dried under conventional conditions. Frozen or dried culture concentrates of Lactobacillus plantarum are commercially available or can be prepared in a manner as described above for L. casei. If a commercial preparation is obtained, it should be mildly disrupted as is the case with L. casei and L. lactis so that the L. plantarum component contains about 23% disrupted cells. The disruption causes the cells to lyse more rapidly and provides more opportunity for enzyme substrate reactions to occur. Our preferred strain of L. plantarum is ATCC 39542. Kid lipase and calf lipase are commercially available. Other animal, plant or microbial lipases which have a similar specificity may also be used. Suitable specificity for a particular lipase or combination of lipases can be determined by inoculating varying levels of lipolytic enzyme into a fresh cheddar cheese slurry containing- 50"-550C moisture. This slurry can be incubated at elevated temperatures ±300-370C) for varying lengths of time. Organoleptic evaluation must be made on a daily basis to determine acceptability of the free fatty acids balance relative to an aged cheddar cheese balance. As an example, a small quantity of slurry is removed for evaluation periodically throughout the incubation period. Early in the flavor development period, the slurried material can be graded directly without dilution. As the free fatty acid flavor intensifies with time, the slurry can be diluted in low fat milk at 6.0% for grading. This bland milk medium aids as a carrier for improved sensory judgments. These lipases, while known to be useful in cheese manufacture, work synergistically with the particular lactobacillus microorganisms described herein to provide the desired curing of American cheese so as to provide the desired flavor in a shorter time than would be the case without the addition of the fast curing ingredients. An optional fourth ingredient in the preparation of the present invention is a microbial, neutral protease having specific activity of 650,000 PU/g and used in an amount of from zero to 600,000 protease activity units (PU) per 454 kg of milk as determined by the standard protease assay. Suitable proteases can be obtained from BaciZZus subtigus, B. amyZo- liquefacians and AspergilZus oryzae. The known methods of making American cheese involve adding milk (adjusted to a temperature of from 86"F to 900F) to the cheesemaking vat. A starter comprising a lactic acid producing bacteria is added to the milk and stirred as the milk ripens due to the development of acidity through bacterial action. The preparation for accelerated aging described herein is preferably added to the cheese milk before the addition of a coagulant but can also be added to the curd with salt before pressing. Rennet (either animal or microbial in origin) is then mixed thoroughly into the ripened milk and the stirring is stopped whereupon the milk is permitted to sit so as to form a coagulum or curd. When the curd is sufficiently firm, it is cut into cubes which are stirred continuously as the whey is expelled during which time the curd is usually heated over a period of about 1/2 hour to a temperature of about 1020F. After draining the whey from the curd, the curd is packed several inches deep, and when the curd is firm enough to be turned without breaking, it is cheddared, that is cut into slabs about 7 inches wide, and turned frequently. After cheddaring, the curd is milled and stirred while salt is added. This causes additional whey to be expelled. After salting and draining, the curd is placed in cloth-lined metal hoops and pressed for several hours to form a cheese. After pressing, the cheese is removed from the hoop and wrapped in a suitable material for curing and aging. Curing usually requires at least 60 days to provide what is known as young or low-flavored cheese. Lengthier curing periods of from about 3 to 6 months provide mildly flavored cheddar cheese. Aged cheddar cheese is produced by curing for 6 to 12 months or longer in some cases. The lengthy time required for aging results in very substantial space requirements to produce and store large quantities of cheese. Thus, the economic advantages of accelerating the aging process are apparent. The method of practicing the present invention is further illustrated by the following examples: Example I To 40,000 pound quantities of cheese milk the following ingredients were added as it was added to a cheese vat: 1) 0.8% fully-ripened bulk starter containing a high population of S. Cremoris and S. Lactus. 2) 1589.0 g of dried L. casei or L. lactis having an activity of 480 protease units/g (67% dis rupted cells). 3) 164.0 g of dried L. plantarum providing-a count to the milk of 5.0 x 106 CFU/ml as based on the original culture concentrate count (23% dis rupted cells). 4) powdered calf lipase (6.26 g) from a stock preparation of 2,500 RU/g. 5) kid lipase (13.12 g) from a stock source having 5,500 RU/g. The preceding mixture was allowed to ripen for 1 hour after which 328 ml of single-strength calf rennet was added. The proper coagulum was formed within 30 minutes at which point cutting occurred. Standard cheesemaking procedures were followed to provide a cheddared curd which was milled when a pH of 5.40 was reached. The milled curd was salted to provide a finished product containing 1.8% (w/w) salt. The finished hooped curd had a final moisture content of 37.5% and fat content of 33.0t. The blocks were shrink film wrapped and aged at 50"F for 3 months which results in the production of a typical aged cheddar cheese flavor. Longer aging at 500F results in strongly aged cheddar cheese flavors which simulate 12 month flavors with only 6 month storage. The flavor development rate can be substantially reduced by lowering the aging temperature from 50"F to 400F or below. Example II To 40,000 pound quantities of cheese milk the following ingredients were added as the milk was added to a cheese vat: 1) 0.8% fully-ripened bulk starter containing a high population of S. Cremoris and S. Cactus. 2) 1589.0 g dried L. casei or L. Zactis having an activity of 480 protease units/g. 3) dried L. piantarurn providing a count to the milk of 5.0 x 106 CFU/ml as based on the original culture concentrate count. 4) powdered calf lipase (6.26 g) from a stock pre paration of 2,500 RU/g. 5) kid lipase (13.12 g) from a stock source of lipase having 5,500 RU/g activity. 6) 10.0 g of protease from Bacillus amyZoZiquefacieus (dry powder having an activity of 650,000 PU/g). The mixtures of special starters, enzymes and milk are allowed to ripen for 1 hour after which 93.7 g rennet powder (315 ó) is added for clotting purposes. The desired coagulum was formed within 30 minutes at which time cutting took place. Following standard cheesemaking procedures, the cheddared curd was milled when a pH of 5.40 was reached. The milled curd was salted to give a finished product of 1.88 and the finished hoop curd had a final moisture content of 37.58 and fat of 33.0t. The blocks were wrapped in shrink film and aged at 50"F for 3 months during which time a typical aged cheddar cheese flavor is produced. This product can be further aged at 50OF resulting in strongly aged cheddar cheese flavors simulating 12 month flavors with only 6 month storage. The flavor development rate can be substantially reduced by lowering the aging temperature from 500F to 400F or below. While a curing temperature of 50"F is preferred, a temperature range of 36 to 600F is suitable. In addition to accelerating the aging process, the preparation of the present invention has been found to prevent the development of defects in the cheese during the aging process. It has been observed in the typical day to day manufacturing of cheddar cheese that there are many variables which will influence the development of off-flavors. In the numerous field trials associated with this ripening system, the control vat of cheese on occasion has produced bitter, sulfur and/or out of balance, fruity, rancid flavor defects. These defects may be recognized at low levels after 1 to 2 months aging or, in some cases, after 3 to 6 months. Generally once these off-flavors are observed, they intensify very quickly. The experimental vats produced on the same day using the identical ingredients and cheesemaking procedures as the control, scored very well in flavor evaluation. Specific trials have been conducted where strongly bitter starter cultures have been intent ion- ally used which resulted in bitter cheese within 2 months. Those vats containing the accelerated ripening system of the present invention have not produced this defect even when aged for 6 months and beyond.

Claims: CLAIMS: 1. A composition for accelerating the aging process of cheese which comprises: a) a preserved, partially disrupted preparation of the lactic acid bacteria Lactobacillus casei, L. lactis or a mixture thereof; b) a preserved culture concentrate of partially disrupted cells of Lactobacillus plantarum; and c) a blend of dried pre-gastic lipases. 2. The composition of claim 1 where in the preserved lactic acid bacteria is dried L. casei. 3. The composition of claim 1 wherein the preserved lactic acid bacteria is dried L. lactis. 4. The composition of claim 3 wherein the L. lactis is designated as ATCC 39538. 5. The composition of claim 1 wherein the preserved L. plantarum is designated as ATCC 39542 and is in the dried form. 6. The composition of claim I wherein there is inclu ded a microbial neutral protease having a specific activity of 650,000 PU/g and is present in an amount of sufficient to provide up to 600,000 protease activity units per 45 kg of cheese milk. 7. The composition of claim 1 wherein the pregastric lipases are calf or kid lipases. 8. A method for accelerating the aging process of American-type cheese which comprises adding to the cheese during its manufacture a composition which comprises: a) a preserved, partially disrupted preparation of the lactic acid bacteria Lactobacillus casei or L. lactis or a mixture thereof; b) a preserved culture concentrate of partially disrupted cells of Lactobacillus plantarum; and c) a blend of dried pre-gastric lipases. 9. The method of claim 8 wherein the preserved lactic acid bacteria is dried L. casei. 10. The method of claim 8 wherein the preserved lactic acid bacteria is dried L. lactis. 11. The method of claim 10 wherein the L. lactis is ATCC 39538. 12. The method of claim 8 wherein the preserved L. planta rum is designated as ATCC 39542 and is in the dried form. 13. The method of claim 8 wherein the L. casei, L. lac tis or mixture thereof is present in an amount of from 0.001 to 0.05 weight percent of milk used in the cheese manufacture based on an activity of 140 standard protease units per gram of preser ved lactic acid bacteria. 14. The method of claim 13 wherein the amount is from 0.025 to 0.03 weight percent. 15. The method of claim 8 wherein the L. plantarum is present in an amount of from 1 x 104 to 5 x 107 CFU per milliliter of milk used in the cheese manu facture based on the cell population before disrup tion. 16. The method of claim 8 wherein the composition added to the cheese includes a microbial neutral protease having a specific activity of 650.000 PU/g in an amount sufficient to provide up to 600,000 protease activity units per 454 kg of milk used in the cheese manufacture. 17. The method of claim 8 wherein the pre-gastric lipase is kid lipase and is used in an amount equivalent to 0.015 to 1.0 gram per 454 kg of milk used in the cheese manufacture based on the kid lipase having an activity of 2,500 Ramsey Units (RU) per gram. 18. The method of claim 17 wherein the amount of kid lipase is from 0.30 to 0.35 g/454 kg of milk. 19. The method of claim 9 wherein the pre-gastric lipase is calf lipase and is used in an amount equivalent to 0.0075 to 0.5 gram per 454 kg of milk used in the cheese manufacture based on the calf lipase having an activity of 5,500 Ramsey Units (RU) per gram. 20. The method of claim 19 wherein the amount of calf lipase is from 0.13 to 0.18 9/454 kg of milk.

Inventors: BARACH, JEFFREY T., TALBOTT, LARRY L.
Applicants: MILES LAB
Application year: 1985
Main classification: A23C19
IPC codes: A23C19