Classifications

• • A—HUMAN NECESSITIES
  • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
  • A23C—DAIRY PRODUCTS, e.g. MILK, BUTTER OR CHEESE; MILK OR CHEESE SUBSTITUTES; MAKING THEREOF
  • A23C19/00—Cheese; Cheese preparations; Making thereof
  • A23C19/06—Treating cheese curd after whey separation; Products obtained thereby
  • A23C19/068—Particular types of cheese
  • A23C19/0682—Mould-ripened or bacterial surface ripened cheeses
• • A—HUMAN NECESSITIES
  • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
  • A23C—DAIRY PRODUCTS, e.g. MILK, BUTTER OR CHEESE; MILK OR CHEESE SUBSTITUTES; MAKING THEREOF
  • A23C19/00—Cheese; Cheese preparations; Making thereof
  • A23C19/02—Making cheese curd
  • A23C19/032—Making cheese curd characterised by the use of specific microorganisms, or enzymes of microbial origin
  • A23C19/0323—Making cheese curd characterised by the use of specific microorganisms, or enzymes of microbial origin using only lactic acid bacteria, e.g. Pediococcus and Leuconostoc species; Bifidobacteria; Microbial starters in general
• • A—HUMAN NECESSITIES
  • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
  • A23C—DAIRY PRODUCTS, e.g. MILK, BUTTER OR CHEESE; MILK OR CHEESE SUBSTITUTES; MAKING THEREOF
  • A23C19/00—Cheese; Cheese preparations; Making thereof
  • A23C19/02—Making cheese curd
  • A23C19/05—Treating milk before coagulation; Separating whey from curd

Definitions

• the invention relates to a method for treatment of milk which is to be used in the preparation of cheese, the method comprises adding a bacterial culture to the milk. It further relates to the resulting milk, and to the milk's use in a cheese process.
• Raw milk received for cheese production has to be stored until it can be used for cheese production, mainly due to bottlenecks in the cheese plant.
• the mineral balance of the milk is displaced, minerals lost, and it therefore looses some of its original ability to coagulate and undergo syneresis, two very important properties in cheese making (Lane, C. N., Sousa, MJ., and McSweeney, P.L.H. (2001)).
• the milk In order to restore these properties to the milk, especially milk to be used for soft cheese such as camembert, the milk normally undergoes a processing, a so-called ⁇ cold-maturation" step, which purpose is to prepare the milk for cheese making.
• Cold maturation consists of physical and biological maturation that aims at obtaining five objectives, generally believed to make the milk more suitable for cheese making (Pernoud S. and Mayer H. L (2008)):
• the milk In cold maturation the milk normally undergoes a mild heat treatment (thermisation, e.g. 62 °C for 20 seconds) or pasteurisation (e.g. 72C for 15 seconds) to remove psycrotroph bacteria such as Listeria species. CaCI2 is added and the milk is kept at 10-15 0 C for 14 to 18 hrs to restore the calcium balance of the milk (physical maturation).
• the milk is added a lactic acid bacterial culture to achieve biological maturation as described above. Normally, lactic acid bacteria cultures that acidify milk well are used to obtain biological maturation as described above.
• the milk is normally pasteurised (e.g., 72 0 C for 20 seconds) to kill and lyse the culture used for the biological maturation thereby releasing bacterial enzymes that may assist in ripening.
• the pH of renneting is to a wide extent governed by the pH reached in cold maturation. It is well known, that the level of pH during renneting will have an important influence on coagulation and syneresis and therefore the moisture content and yield of soft cheese (Mietton, B., Gaucheron, F. and Sala ⁇ n-Michel, F. (2004)).
• the moisture content of the soft cheese has a direct influence on yield and texture and an indirect influence on ripening.
• lactic acid bacteria cultures that only acidify milk slowly are less sensitive to variations in process temperature and process times during cold maturation. And that this may result in a cheese product with uniform yield and quality from batch to batch, ensuring more profitable cheese making.
• Experiments wherein a protease negative culture of lactic acid bacteria has been added to milk have surprisingly revealed that the resulting milk results in standardized milks which can be used for cheese production, and the present inventors contemplate that addition to milk of other slow acidifying lactic acid bacteria will result in a standardized milk.
• the present inventors have invented a novel method for maturation and storage of the milk to be used for cheese production, the method results in more standardized milk for cheese making, which may result in a cheese product with uniform yield and quality from batch to batch.
• the method comprises addition of a slow acidifying lactic acid bacteria culture that exhibits limited acidification in the milk.
• the cold maturation acidification of the milk by the culture is less sensitive to fluctuations in process temperatures, which may reduce batch to batch variations in a cheese product.
• the acidification of the milk by the culture is less sensitive to fluctuations in process times, which may reduce batch to batch variations in a cheese product.
• the present invention relates to a method of treatment of milk (e.g. to be used for production of cheese), said method comprises adding to the milk a culture of bacteria which: is not able to acidify the milk, or acidifies the milk slowly, and/or is not able to grow in milk, or grows slowly in milk.
• the invention also relates to a method of treatment of milk (e.g. to be used for production of cheese), said method comprises adding to the milk a culture of bacteria which are (substantially) protease negative and/or (substantially) incapable of degrading milk proteins (e.g. casein).
• the present invention further relates to a method of treatment of milk (e.g. to be used for production of cheese), said method comprises adding to the milk a culture of bacteria, which culture is characterized by that it lowers the pH of the milk less than 0.25 pH units per hour at 30 degrees C, such as less than 0.20 pH units, less than 0.15 pH units, or less than 0.10 pH units, when inoculated at a quantity of 10E6 cfu (cell forming units) per ml milk (esp RSM or laboratory milk).
• a culture of bacteria which culture is characterized by that it lowers the pH of the milk less than 0.25 pH units per hour at 30 degrees C, such as less than 0.20 pH units, less than 0.15 pH units, or less than 0.10 pH units, when inoculated at a quantity of 10E6 cfu (cell forming units) per ml milk (esp RSM or laboratory milk).
• the bacteria (in the culture) are selected from the group consisting of: bacteria which are (substantially) protease negative, - bacteria which are (substantially) incapable of degrading milk proteins (e.g. casein), bacteria which are (substantially) incapable of degrading lactose, lactate dehydrogenase (Ldh) defective bacteria (cf. EP0928333B1) bacteria which are thymidine auxotrophic mutants (thyA) (cf. EP1102837B1), and bacteria which are defective in pyruvate formate-lyase (PfI) (cf. EP0928333B1).
• bacteria which are (substantially) protease negative e.g. casein
• bacteria which are (substantially) incapable of degrading lactose e.g. casein
• lactate dehydrogenase (Ldh) defective bacteria cf. EP0928333B1
• the culture may comprise one or more strains (or bacteria belonging to such a strain) of the group defined above, optionally together with other strains, as long as the culture as a whole is not able to grow in milk, grows slowly in milk, or is (substantially) protease negative, or is (substantially) incapable of degrading milk proteins, lactose, etc..
• the culture of bacteria is a culture of lactic acid bacteria, such as a culture of one or more strains, selected from the group consisting of Lactococcus spp., Streptococcus spp., Lactobacillus spp., Leuconostoc spp., Pseudoleuconostoc spp., Pediococcus spp., Brevibacte ⁇ um spp., Enterococcus spp. and Propionibacterium spp.
• the culture may comprise one or more strains of other species than these.
• the culture should lower the pH of the milk less than 0.25 pH units per hour at 30 degrees C, such as less than 0.20 pH units, less than 0.15 pH units, or less than 0.10 pH units, when inoculated at a quantity of 10E6 cfu (cell forming units) per ml milk (esp. in RSM or in laboratory milk).
• a culture of a strain selected from the group consisting of DN224 (DSM11037), DIM223 (DSM11036), DN221 (DSM11034), DIN227 (DSM11040), MBP71 (DSM12891), DN105 (DSM12289), or mutants or variants of any of these strains.
• the method of the invention further comprises: a) no microorganisms, which are able to degrade (a substantial part of) milk proteins, are added to the milk, or are allowed to act on the milk proteins; and/or b) the milk is not subjected to any (substantial) degradation of milk proteins.
• the present invention relates to a method of treatment of milk, said method comprises adding to the milk a culture of bacteria which are (substantially) protease negative and/or (substantially) incapable of degrading milk proteins (e.g. casein).
• milk is treated in a way such that less than 30 % (such as less than 20%, less than 10%, or less than 5%) of the milk proteins (e.g. casein) are degraded, or less than 30% % (such as less than 20%, less than 10%, or less than 5%) of the lactose is degraded, such as when in the milk is stored 24 hours at 15 degrees C after inoculation with 10E6 cfu/ml milk.
• the milk proteins e.g. casein
• 30% % such as less than 20%, less than 10%, or less than 5%
• the method may further comprise: a) heat treatment of the milk (such as to a temperature in the range 40-80 degrees C, e.g. pasteurization or thermisation); b) cooling the milk (such as to a temperature in the range 5-15 degrees C); and c) storing the milk (such as for 1-48 hours); and d) optionally heat treatment of the milk (e.g. pasteurization or thermisation).
• the culture should added after step a).
• the milk is kept at a temperature below 20 degrees C, such as at a temperature in the range 5-20 degrees C, or in the range 5-15 degrees C, such as from 1-48 hours (e.g. from 4-24 hours or from 5-20 hours). It is presently preferred that the culture is added to the milk in a final concentration of 10E3 to 10E12 CFU pr ml milk, such as from 10E5 to 10E10 cfu/ml, or from 10E7 to 10E9 cfu/ml.
• the invention relates to a milk, e.g. for use in the production of cheese, which is obtainable by a method of the invention, or a milk which is treated using a method of the invention.
• the invention relates to a method for preparing cheese, wherein a milk of the present invention or a milk obtainable by a method of the invention, is contacted with a a) a culture of lactic acid bacteria, such as a culture of protease positive bacteria; and b) a coagulant, such as a protease (e.g. rennet, a chymosin or a microbial coagulant).
• a a culture of lactic acid bacteria such as a culture of protease positive bacteria
• a coagulant such as a protease (e.g. rennet, a chymosin or a microbial coagulant).
• the method may comprise further cheese making steps. Such steps are known to the person skilled in the art.
• the invention relates to a cheese obtainable by the method of the invention, such as a soft cheese, e.g. camembert.
• milk refers to the lacteal secretion obtained by milking any mammal, such as cows, sheep, goats, buffaloes or camels.
• the milk is cow's milk, and especially raw cow's milk.
• milk also comprises compositions comprising milk, and milk compositions that have been treated, e.g. chemically, enzymatically, and/or mechanically.
• microorganism may include any bacterium, or fungus being able to ferment the milk substrate. Lactic acid bacteria are preferred microorganisms.
• lactic acid bacteria designates a gram-positive, microaerophilic or anaerobic bacterium, which ferments sugars with the production of acids including lactic acid as the predominantly produced acid, acetic acid and propionic acid.
• lactic acid bacteria are found within the order "Lactobacillales" which includes Lactococcus spp., Streptococcus spp., Lactobacillus spp., Leuconostoc spp., Pseudoleuconostoc spp., Pediococcus spp., Brevibacterium spp., Enterococcus spp. and Propionibacterium spp. Additionally, lactic acid producing bacteria belonging to the group of the strict anaerobic bacteria, bifidobacteria, i.e. Bifidobacterium spp., are generally included in the group of lactic acid bacteria.
• a strain which might be used in the present invention is a strain which has lost the capability of de novo synthesising essentially compounds (also referred to in the art as an "auxotrophic strain"). Therefore, in preferred embodiments, the bacterial strain is a strain being auxothrophic in respect of a compound which is not present in the milk and which is required by the strain for growth.
• culture refers to any sample or item that contains one or more microorganisms.
• "Pure cultures” are cultures in which the organisms present are only of one strain of a particular genus and species. This is in contrast to “mixed cultures,” which are cultures in which more than one genus and/or species of microorganism are present. In some embodiments of the present invention, pure cultures find use, but normally a culture as used in present invention contains more than one strain.
• laboratory milk is a reconstituted skim milk (RSM) with 9.5% dry-matter on a weight basis that has been subjected to temperatures of 99C for 30 minutes before use.
• a “slow acidifying" bacteria culture (or a culture which acidifies the milk slowly) is a culture which has a maximum rate of acidification of 0.25 pH units per hour at 30 degrees C when inoculated at a quantity of 10 ⁇ 6 cfu (10E6 cell forming units) per ml laboratory milk.
• the culture as a whole should have the max rate of acidification of 0.25 pH per hour at 3OC as when inoculated 10E6 cfu/ml milk as defined above).
• any other culture than a slow acidifying culture is defined as a "fast acidifying" culture.
• the term “grows slowly” in the present context means that the cell count in the milk doubles in more than two hours (such as more than 4, 6, 10, or 24 hours) at 30 degrees C when inoculated at a quantity of 10 A 6 cfu (10E6 cell forming units) per ml (laboratory) milk.
• the enzymatic activity is less than 10% of the type strain of the same species or of the mother strain from which the mutant is obtained, measured under identical conditions.
• Soft cheese is defined as any Rennet coagulated cheese that contains about 70 - 74 % moisture on a non fat solids basis and is produced without scalding and pressing.
• mutant should be understood as a strain derived, or a strain which can be derived, from a strain of the invention (or the mother strain) by means of e.g. genetic engineering, radiation and/or chemical treatment. It is preferred that the mutant is a functionally equivalent mutant, e.g. a mutant that has substantially the same, or improved, properties (e.g. regarding acidification speed) as the mother strain. Such a mutant is a part of the present invention.
• mutant refers to a strain obtained by subjecting a strain of the invention to any conventionally used mutagenization treatment including treatment with a chemical mutagen such as ethane methane sulphonate (EMS) or N- methyl-N'-nitro-N-nitroguanidine (NTG), UV light, or to a spontaneously occurring mutant.
• a mutant may have been subjected to several mutagenization treatments (a single treatment should be understood one mutagenization step followed by a screening/selection step), but it is presently preferred that no more than 20, or no more than 10, or no more than 5, treatments (or screening/selection steps) are carried out.
• a presently preferred mutant less that 5%, or less than 1% or even less than 0.1% of the nucleotides in the bacterial genome have been shifted with another nucleotide, or deleted, compared to the mother strain.
• variant should be understood as a strain which is functionally equivalent to a strain of the invention, e.g. having substantially the same, or improved, properties e.g. regarding acidification speed). Such variants, which may be identified using appropriate screening techniques, are a part of the present invention.
• Example 1 reduced sensitivity towards process delays in a laboratory system
• F-DVS Free Direct Vat Set
• Culture A Bulk Item 699759 F-DVS SICO-Ol-O-PRT- and Culture B: Bulk Item 693091 F-DVS SICO-14-O were Inoculated 10 g per 100 kg milk in re-constituted skim milk (9.5 % DM) and heat treated milk (99C for 30 minutes), cooled to fermentation temperature 1OC, 15C and 2OC, respectively.
• Culture A represents the slow acidifying culture.
• Culture B represents the fast acidifying culture.
• F-DVS frozen direct vat set
• Culture A represents the slow acidifying culture.
• Culture B represents the normal acidifying culture.
• FIG. 2 shows the pH after 12 hours at 1OC, 15C, and 2OC, respectively.
• the horizontal line represents pH 6.4 our target for soft cheese.
• the blue line is a calculated fit to the data.
• pH 6.4 If pH 6.4 is sought, it will be reached in 12 hours at 14 0 C with culture B (filled Triangles), and in 12 hours at 15.5 °C with culture A (Filled Circles).
• a variation in temperature of 13 to 15 °C will translate into a pH variation of about 0.3 pH units for culture B.
• a variation in temperature of 14.5 to 16.5 0 C will translate into a pH variation of about 0.125 pH units for culture A.
• Example 3 reduced batch to batch variation due to process delays in soft cheese Culture A: slow acidification culture 699759 F-DVS SICO-Ol-O-PRT-, Culture B: traditional fast acidifying cold maturation culture 501691 F-DVS FLORA-DANICA, both obtainable from Chr Hansen, H ⁇ rsholm, Denmark
• Pasteurized milk is inocultated with the culture (72C for 15s), cooled to 15C with 5 g culture per 100 liter milk, and fermented at 15C for until pH 6.4 is reached, note the time (TO) needed.
• the other half of the milk is allowed to continue the fermentation at TO + 1 hr, then pasteurized (72C for 15s) the milk and to produce camembert according to a standard soft cheese make (see below).
• the experiment is repeated a sufficient number of times, e.g. six times for each culture.
• the weight, moisture, and flavor difference between the cheeses made from TO milk and those from TO+lhr milk is calculated.
• Example 4 reduced batch to batch variation due to temperature differences in soft cheese Culture A: slow acidification culture 699759 F-DVS SICO-Ol-O-PRT-, Culture B: traditional fast acidifying cold maturation culture 501691 F-DVS FLORA-DANICA, both obtainable from Chr Hansen, H ⁇ rsholm, Denmark.
• a and B Inoculate simultaneously with 5 g culture per 100 liter pasteurized milk (72C for 15s), cooled to 14, 15, and 16C, respectively. Ferment until pH 6.4 is reached at 15C, and pasteurize (72C for 15s) each batch of milk simultaneously.
• Example 5 a slow and a fast acidifying culture
• F-DVS Free Direct Vat Set
• Culture A (solid bold line) represents the slow acidifying culture.
• Culture B (dashed bold line) represents the fast acidifying culture.
• the thin dotted line represents the maximum acidification rate of 0.25 pH units per hour (i.e. the maximum slope of the acidification curve) defining a slow acidifying culture in the context of this invention.

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• Life Sciences & Earth Sciences (AREA)
• Chemical & Material Sciences (AREA)
• Engineering & Computer Science (AREA)
• Food Science & Technology (AREA)
• Polymers & Plastics (AREA)
• Microbiology (AREA)
• Dairy Products (AREA)

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• 2010
  • 2010-06-04 WO PCT/DK2010/000091 patent/WO2010139333A1/en active Application Filing
  • 2010-06-04 EP EP10727343A patent/EP2437613A1/de not_active Withdrawn

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