CN104894036A - Cheese starter and preparation method and application thereof - Google Patents
Cheese starter and preparation method and application thereof Download PDFInfo
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- CN104894036A CN104894036A CN201510378605.9A CN201510378605A CN104894036A CN 104894036 A CN104894036 A CN 104894036A CN 201510378605 A CN201510378605 A CN 201510378605A CN 104894036 A CN104894036 A CN 104894036A
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- 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
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Abstract
The invention discloses a cheese starter and a preparation method and application thereof. The cheese starter comprises Lactococcus lactis subsp.lactis CGMCC No.10752, Lactococcus lactis subsp.lactis CGMCC No.10748, Lactococcus lactis subsp.lactis CGMCC No.10751 and Leuconostoc mesenteroides CGMCC No.10750, wherein the viable count ratio of Lactococcus Lactis subsp.lactis CGMCC No.10752 to Lactococcus lactis subsp.lactis CGMCC No.10748 to Lactococcus lactis subsp.lactis CGMCC No.10751 to Leuconostoc mesenteroides CGMCC No.10750 is (0.5-1):(1-1.5):(1-2):(1-2). The cheese starter can be used for processing cheese by using a good alternate effect between starter strains, and the preparation method is simple, convenient and feasible.
Description
Technical field
The present invention relates to milk-product field, be specifically related to a kind of cheese starter and its preparation method and application.
Background technology
Cheese, has another name called cheese, has taste miscellaneous, mouthfeel and form.Cheese take milk as raw material, containing rich in protein and lipid, has very high nutritive value.Along with our people's growth in the living standard, the demand of cheese is got more and more.In cheesemaking and ripening process, microorganism species plays an important role, and facilitates the matter structure of product and the formation of local flavor.The formation of cheese starter to cheese matter structure and typical flavor of cheese plays extremely important effect.Cheese starter produces acid in cheese manufacturing process, can improve the activity of rennin, contributes to getting rid of whey, suppressing harmful bacterial growth, and produces the formation that various enzyme participates in cheese typical flavor and matter structure.
In the screening process of cheese milk acid bacterium, because the Microflora of screening is many, and in order to reach the Effective selection to good characteristic bacterial strain, needing optimal screening step, is generally that the bacterial classification choosing excellent performance combinationally uses by screening large batch of bacterium in the world, by 2-6 strain bacterium being combined, to make up the defect of single strain in product acid, product perfume etc., the necessary symbiosis of these bacterium is good simultaneously, could be used for being developed to cheese starter.Due to cheese with the screening of bacterium, combine, prepare the ins and outs such as starter all grasp by external business bacterial classification company.How agriculture products, defining method, filters out excellent bacterial classification, how to combine bacterial strain, be developed to excellent cheese starter and be substantially in blank at home.
Vast territory and abundant resources in China, and microorganism resource is enriched, and characteristic milk-product are also extremely abundant, therefrom filter out the bacterial strain of applicable cheese, and are combined into starter, may be used for developing sinicization cheese.
Summary of the invention
Technical problem to be solved by this invention is, for lack at present be used for cheesemaking addicted to warm nature starter, be difficult to combine simultaneously by produce acid type starter fast and produce fragrant fermenting agent be applied to cheese making machine standby in deficiency, a kind of cheese starter and its preparation method and application are provided.Described cheese starter can be used in cheese, especially the preparation of Gouda cheese, and taken into account cheese making machine and produced for middle the requirement that acid is fast and product is fragrant, proteolytic activity is strong, and each component balanced result of flavour substances is good.Described preparation method is easy to operation, is convenient to make described cheese starter be applied to industrialization and prepares on a large scale in cheese.
The invention provides a kind of cheese starter, it comprises Lactococcus lactis subsp.lactis (Lactococcus lactissubsp.lactis) CGMCC No.10752 (BD401), Lactococcus lactis subsp.lactis (Lactococcus lactis subsp.lactis) CGMCC No.10749 (BD2263), Lactococcus lactis subsp.lactis (Lactococcus lactis subsp.lactis) CGMCC No.10751 (BD164) and Leuconostoc mesenteroides (Leuconostoc mesenteroides) CGMCC No.10750 (LM79), described Lactococcus lactis subsp.lactis CGMCC No.10752, described Lactococcus lactis subsp.lactis CGMCC No.10749, the viable count ratio of described Lactococcus lactis subsp.lactis CGMCC No.10751 and described Leuconostoc mesenteroides CGMCC No.10750 is (0.5 ~ 1): (1 ~ 1.5): (1 ~ 2): (1 ~ 2).
Preferably, the viable count ratio of described Lactococcus lactis subsp.lactis BD401, described Lactococcus lactis subsp.lactis BD2263, described Lactococcus lactis subsp.lactis BD164 and described Leuconostoc mesenteroides LM79 is 1:1:1:1.
Preferably, described cheese starter also comprises auxiliary material.Described auxiliary material is the auxiliary material of this area routine, for the preparation of cheese starter.Preferably comprise in water, lactose, sucrose, maltodextrin, Sodium Glutamate, gelatin, glycerine, sorbyl alcohol, trehalose, yeast extract paste and beta-cyclodextrin one or more.
The form of described cheese starter is the form of this area routine, be preferably liquid starter, frozen fermented agent or throw type leaven, be more preferably throw type leaven, namely described cheese starter without the need to activating it, the pre-treatment step such as enlarged culturing and directly drop into fermented substrate and use.
The invention provides a kind of preparation method of cheese starter, it comprises the following steps:
(1) Lactococcus lactis subsp.lactis CGMCC No.10752, Lactococcus lactis subsp.lactis CGMCC No.10749, Lactococcus lactis subsp.lactis CGMCC No.10751 and Leuconostoc mesenteroides CGMCC No.10750 are inoculated in substratum, obtain culture;
(2) from the culture of step (1) gained, Lactococcus lactis subsp.lactis CGMCCNo.10752 is obtained, Lactococcus lactis subsp.lactis CGMCC No.10749, Lactococcus lactis subsp.lactis CGMCC No.10751 and Leuconostoc mesenteroides CGMCC No.10750, according to Lactococcus lactis subsp.lactis CGMCC No.10752, Lactococcus lactis subsp.lactis CGMCC No.10749, the viable count of Lactococcus lactis subsp.lactis CGMCC No.10751 and Leuconostoc mesenteroides CGMCC No.10750 is (0.5 ~ 1): (1 ~ 1.5): (1 ~ 2): the ratio mixing of (1 ~ 2).
Step (1) is: be inoculated in substratum by Lactococcus lactis subsp.lactis CGMCC No.10752, Lactococcus lactis subsp.lactis CGMCC No.10749, Lactococcus lactis subsp.lactis CGMCC No.10751 and Leuconostoc mesenteroides CGMCC No.10750, obtains culture.Wherein, preferably, the step of activation is comprised before described inoculation.The algebraically of described activation is the algebraically of this area routine, is preferably 2 ~ 3 generations.The substratum of described activation is the substratum of this area routine, described Lactococcus lactis subsp.lactis BD401, described Lactococcus lactis subsp.lactis BD2263, described Lactococcus lactis subsp.lactis BD164 and described Leuconostoc mesenteroides LM79 can be grown, it is preferably degreasing milk medium, it is more preferably 10% degreasing milk medium, after described degreasing milk medium is dissolved in water by milk powder, mixing obtains, and described per-cent is the mass percent that described milk powder accounts for the total mass of described milk powder and described water.The temperature of described activation is the temperature of this area routine, is preferably 28 ~ 32 DEG C.The time of described activation is the time of this area routine, is preferably 16 ~ 24 hours.Described substratum is the substratum of this area routine, can Lactococcus lactis subsp.lactis BD401, described Lactococcus lactis subsp.lactis BD2263, described Lactococcus lactis subsp.lactis BD164 and described Leuconostoc mesenteroides LM79 described in enlarged culturing, it is preferably degreasing milk medium, be more preferably 10% degreasing milk medium, described per-cent is quality volume percent.The temperature of described cultivation is the temperature of this area routine, is preferably 28 ~ 32 DEG C.The time of described cultivation is the time of this area routine, is preferably 16 ~ 24 hours.The number of times of described cultivation is the number of times of this area routine, is preferably 2 ~ 3 times.
Step (2) is: from the culture of step (1) gained, obtain Lactococcus lactis subsp.lactis CGMCC No.10752, Lactococcus lactis subsp.lactis CGMCC No.10749, Lactococcus lactis subsp.lactis CGMCC No.10751 and Leuconostoc mesenteroides CGMCC No.10750, according to Lactococcus lactis subsp.lactis CGMCC No.10752, Lactococcus lactis subsp.lactis CGMCC No.10749, the viable count of Lactococcus lactis subsp.lactis CGMCC No.10751 and Leuconostoc mesenteroides CGMCC No.10750 is (0.5 ~ 1): (1 ~ 1.5): (1 ~ 2): the ratio mixing of (1 ~ 2).Wherein, preferably, according to the viable count of Lactococcus lactis subsp.lactis CGMCC No.10752, Lactococcus lactis subsp.lactis CGMCC No.10749, Lactococcus lactis subsp.lactis CGMCC No.10751 and Leuconostoc mesenteroides CGMCC No.10750 be the ratio mixing of 1:1:1:1.
Preferably, further comprising the steps of after step (2) completes: to add auxiliary material, mixing.
Wherein, described auxiliary material is the auxiliary material of this area routine, preferably comprises one or more in water, lactose, sucrose, maltodextrin, Sodium Glutamate, gelatin, glycerine, sorbyl alcohol, trehalose, yeast extract paste and beta-cyclodextrin.
The invention provides a kind of described cheese starter and prepare the application in cheese.
Described cheese is the cheese of this area routine, is preferably Gouda cheese, cheddar cheese-and Pa Masen cheese.It is more preferably Gouda cheese.
On the basis meeting this area general knowledge, above-mentioned each optimum condition, can arbitrary combination, obtains the preferred embodiments of the invention.
Agents useful for same of the present invention and raw material are all commercially.
Positive progressive effect of the present invention is: cheese starter of the present invention can prepare cheese, and the cheese obtained by it is very nearly the same, satisfactory with the cheese obtained by commercial fermentation agent in sensory evaluation, matter structure etc.Cheese starter simultaneously (just selects the lactococcal strain that 8 strains are adapted to cheese ripening environment by selecting suitable fermentation strain from 41 strain Lactococcus lactis and 47 strain Leuconostoc mesenteroides bacterial strains, take into account the symbiosis with Leuconostoc mesenteroides and other lactococcal strains, have finally chosen the involved bacterial strain in described cheese agent), and the proportionlity between fermentation strain, properly solve in prior art the deficiency being difficult to combine and producing acid type starter fast and produce fragrant fermenting agent, utilize mutuality good between fermentation strain simultaneously, described cheese starter is made to can be used as the starter of process cheese.In addition, described preparation method is simple and easy to do, can produce cheese starter in batches on a large scale, thus cheese starter is applied in the actual production of milk-product.
biomaterial preservation information
Lactococcus lactis subsp.lactis BD164 of the present invention, be deposited in China Committee for Culture Collection of Microorganisms's common micro-organisms center (CGMCC) on April 27th, 2015, preservation address: No. 3, Yard 1, BeiChen xi Road, Chaoyang District, Beijing City, postcode: 100101, deposit number is: CGMCC No.10751, culture title is BD164, and Classification And Nomenclature is Lactococcus lactis subsp.lactis Lactococcus lactis subsp.lactis.
Lactococcus lactis subsp.lactis BD401 of the present invention, be deposited in China Committee for Culture Collection of Microorganisms's common micro-organisms center (CGMCC) on April 27th, 2015, preservation address: No. 3, Yard 1, BeiChen xi Road, Chaoyang District, Beijing City, postcode: 100101, deposit number is: CGMCC No.10752, culture title is BD401, and Classification And Nomenclature is Lactococcus lactis subsp.lactis Lactococcus lactis subsp.lactis.
Lactococcus lactis subsp.lactis BD2263 of the present invention, be deposited in China Committee for Culture Collection of Microorganisms's common micro-organisms center (CGMCC) April 27 in 2015, preservation address: No. 3, Yard 1, BeiChen xi Road, Chaoyang District, Beijing City, postcode: 100101, deposit number is: CGMCC No.10749, culture title is BD2263, and Classification And Nomenclature is Lactococcus lactis subsp.lactis Lactococcus lactis subsp.lactis.
Leuconostoc mesenteroides LM79 of the present invention, be deposited in China Committee for Culture Collection of Microorganisms's common micro-organisms center (CGMCC) on April 27th, 2015, preservation address: No. 3, Yard 1, BeiChen xi Road, Chaoyang District, Beijing City, postcode: 100101, deposit number is: CGMCC No.10750, culture title is LM79 Leuconostoc mesenteroides, and Classification And Nomenclature is Leuconostoc mesenteroides.
Accompanying drawing explanation
Fig. 1 is the SDS-PAGE collection of illustrative plates of pH4.6 water-insoluble protein, and wherein contrast is control group, and 2 is embodiment 7, is labeled as standard Marker, and α-CN and β-CN represents α-CN albumen and the band corresponding to β-CN albumen respectively.
Embodiment
Mode below by embodiment further illustrates the present invention, but does not therefore limit the present invention among described scope of embodiments.The experimental technique of unreceipted actual conditions in the following example, conventionally and condition, or selects according to catalogue.
The preparation of embodiment 1 cheese starter
(1) by Lactococcus lactis subsp.lactis BD401, Lactococcus lactis subsp.lactis BD2263, Lactococcus lactis subsp.lactis BD164 and Leuconostoc mesenteroides LM79 is inoculated in the degreasing milk medium of 10% (w/v) sterilizing by the inoculum size of 2% (v/v), and (skimmed milk powder is purchased from New Zealand Westland cooperation dairy industry company limited, skimmed milk powder is dissolved in the water and mixes and obtain degreasing milk medium, described per-cent is the mass percent that described skimmed milk powder accounts for described skimmed milk powder and described water total mass) in, be placed in 30 DEG C of constant incubators and cultivate 24 hours, activated for 2 generations, the bacterial strain that must activate.The bacterial strain of activation is inoculated in the degreasing milk medium of 10% (w/v) sterilizing by the inoculum size of 2% (v/v), is placed in 30 DEG C of constant incubators and cultivates and carry out enlarged culturing in 24 hours, repeat 3 times, acquisition culture.
(2) culture of step (1) gained is obtained Lactococcus lactis subsp.lactis BD401, Lactococcus lactis subsp.lactis BD2263, Lactococcus lactis subsp.lactis BD164 and Leuconostoc mesenteroides LM79, then be the ratio mixing of 1:1:1:1 according to the viable count of described Lactococcus lactis subsp.lactis BD401, described Lactococcus lactis subsp.lactis BD2263, described Lactococcus lactis subsp.lactis BD164 and described Leuconostoc mesenteroides LM79, obtain cheese starter.
The preparation of embodiment 2 cheese starter
(1) Lactococcus lactis subsp.lactis BD401, Lactococcus lactis subsp.lactis BD2263, Lactococcus lactis subsp.lactis BD164 and Leuconostoc mesenteroides LM79 are inoculated in the degreasing milk medium (purchased from New Zealand Westland cooperation dairy industry company limited) of 10% (w/v) sterilizing by the inoculum size of 2% (v/v), be placed in 28 DEG C of constant incubators and cultivate 24 hours, activated for 3 generations, obtain the bacterial strain of activation.The bacterial strain of activation is inoculated in the degreasing milk medium of 10% (w/v) sterilizing by the inoculum size of 2% (v/v), is placed in 28 DEG C of constant incubators and cultivates and carry out enlarged culturing in 24 hours, repeat 2 times, acquisition culture.
(2) culture of step (1) gained is obtained Lactococcus lactis subsp.lactis BD401, Lactococcus lactis subsp.lactis BD2263, Lactococcus lactis subsp.lactis BD164 and Leuconostoc mesenteroides LM79, then be the ratio mixing of 0.5:1.5:2:2 according to the viable count of described Lactococcus lactis subsp.lactis BD401, described Lactococcus lactis subsp.lactis BD2263, described Lactococcus lactis subsp.lactis BD164 and described Leuconostoc mesenteroides LM79, obtain cheese starter.
The preparation of embodiment 3 cheese starter
(1) Lactococcus lactis subsp.lactis BD401, Lactococcus lactis subsp.lactis BD2263, Lactococcus lactis subsp.lactis BD164 and Leuconostoc mesenteroides LM79 are inoculated in the degreasing milk medium (purchased from New Zealand Westland cooperation dairy industry company limited) of 10% (w/v) sterilizing by the inoculum size of 2% (v/v), be placed in 32 DEG C of constant incubators and cultivate 16 hours, activated for 3 generations, obtain the bacterial strain of activation.The bacterial strain of activation is inoculated in the degreasing milk medium of 10% (w/v) sterilizing by the inoculum size of 2% (v/v), is placed in 32 DEG C of constant incubators and cultivates and carry out enlarged culturing in 16 hours, repeat 3 times, acquisition culture.
(2) culture of step (1) gained is obtained Lactococcus lactis subsp.lactis BD401, Lactococcus lactis subsp.lactis BD2263, Lactococcus lactis subsp.lactis BD164 and Leuconostoc mesenteroides LM79, then be the ratio mixing of 0.5:1:1:2 according to the viable count of described Lactococcus lactis subsp.lactis BD401, described Lactococcus lactis subsp.lactis BD2263, described Lactococcus lactis subsp.lactis BD164 and described Leuconostoc mesenteroides LM79, obtain cheese starter.
The preparation of embodiment 4 cheese starter
(1) Lactococcus lactis subsp.lactis BD401, Lactococcus lactis subsp.lactis BD2263, Lactococcus lactis subsp.lactis BD164 and Leuconostoc mesenteroides LM79 are inoculated in the degreasing milk medium (purchased from New Zealand Westland cooperation dairy industry company limited) of 10% (w/v) sterilizing by the inoculum size of 2% (v/v), be placed in 32 DEG C of constant incubators and cultivate 16 hours, activated for 3 generations, obtain the bacterial strain of activation.The bacterial strain of activation is inoculated in the degreasing milk medium of 10% (w/v) sterilizing by the inoculum size of 2% (v/v), is placed in 32 DEG C of constant incubators and cultivates and carry out enlarged culturing in 16 hours, repeat 3 times, acquisition culture.
(2) culture of step (1) gained is obtained Lactococcus lactis subsp.lactis BD401, Lactococcus lactis subsp.lactis BD2263, Lactococcus lactis subsp.lactis BD164 and Leuconostoc mesenteroides LM79, then be the ratio mixing of 0.5:1:1:1 according to the viable count of described Lactococcus lactis subsp.lactis BD401, described Lactococcus lactis subsp.lactis BD2263, described Lactococcus lactis subsp.lactis BD164 and described Leuconostoc mesenteroides LM79, obtain cheese starter.
Embodiment 5 cheese starter produces Gouda cheese
1) 100kg fresh cow milk (purchased from pasture, Kingsoft) after filtration, adds 0.01% calcium chloride, stirs, and at 72 DEG C, is cooled to 30 DEG C after 15s pasteurize, and described per-cent is mass percent.Cheese starter obtained for embodiment 1 is poured in the fresh cow milk after pasteurize, makes the viable count of the cheese starter in fresh cow milk reach 10
7cfu/mL, slowly stirs 15min.Leave standstill 28 DEG C of fermentations and add 20mg/100L rennin (purchased from Danisco A/S BJ Rep Office) (wherein after pH reduces by 0.1, rennin dissolves with the sterilized water of 10 times of quality before use, stirring 2min makes it be uniformly dissolved), obtain curdled milk after 30min.
2) by step 1) obtained curd cutting becomes the cubical grumeleuse of length of side 5mm, discharges the whey of cumulative volume 35% after slowly stirring after 15min; Point to add 50 DEG C for 2 times, volume is the water of fresh cow milk cumulative volume 25%, stir 20min and reach 35 DEG C to curd temperature.The whey pH continuing to be stirred to discharge is 6.1, whole whey of draining.Afterwards grumeleuse is entered mould, squeezing is shaped, the demoulding is taken out.Working concentration is salt solution 12 DEG C of salt marsh 20h of 20%, and described per-cent is mass percent.Vacuum packaging afterwards, 10 DEG C, humidity 90%RH, ripe 4 weeks, obtain young Gouda cheese.
The young Gouda cheese smooth texture of gained is tight, high resilience, and shell look yellow and with the ripening stage by thin thickening, tangent plane can have irregular aperture, and smell is gentle, has the fragrance of cream fusing, and with the local flavor of slight nut.
Embodiment 6 cheese starter produces Gouda cheese
1) 100kg fresh cow milk (purchased from pasture, Kingsoft) after filtration, adds 0.02% calcium chloride, stirs, and at 72 DEG C, is cooled to 30 DEG C after 15s pasteurize, and described per-cent is mass percent.Cheese starter obtained for embodiment 1 is poured in the fresh cow milk after pasteurize, makes the viable count of the cheese starter in fresh cow milk reach 10
6cfu/mL, slowly stirs 10min.Leave standstill 35 DEG C of fermentations and add 20mg/100L rennin (purchased from Hansen Corp. of section) (wherein after pH reduces by 0.2, rennin dissolves with the sterilized water of 10 times of quality before use, stirring 2min makes it be uniformly dissolved), obtain curdled milk after 40min.
2) by step 1) obtained curd cutting becomes the cubical grumeleuse of length of side 15mm, discharges the whey of cumulative volume 45% after slowly stirring after 20min; Once add 60 DEG C, volume is the water of fresh cow milk cumulative volume 40%, stir 30min and reach 38 DEG C to curd temperature.The whey pH continuing to be stirred to discharge is 6.1, whole whey of draining.Afterwards grumeleuse is entered mould, squeezing is shaped, the demoulding is taken out.Working concentration is salt solution 15 DEG C of salt marsh 18h of 20%, and described per-cent is mass percent.Vacuum packaging afterwards, 15 DEG C, humidity 90%RH, ripe 17 weeks, obtain ripe Gouda cheese.
Embodiment 7 cheese starter produces Gouda cheese
1) (15% milk powder adds water acquisition 100kg reconstituted milk, described per-cent is mass percent, described milk powder is purchased from bright milk industry) after filtration, add 0.01% calcium chloride, stir, at 72 DEG C, be cooled to 30 DEG C after 15s pasteurize, described per-cent is mass percent.Cheese starter obtained for embodiment 1 is poured in the fresh cow milk after pasteurize, makes the viable count of the cheese starter in fresh cow milk reach 10
7cfu/mL, slowly stirs 15min.Leave standstill 32 DEG C of fermentations and add 20mg/100L rennin (purchased from Danisco A/S BJ Rep Office) (wherein after pH reduces by 0.2, rennin dissolves with the sterilized water of 10 times of quality before use, stirring 2min makes it be uniformly dissolved), obtain curdled milk after 30min.
2) by step 1) obtained curd cutting becomes the cubical grumeleuse of length of side 5mm, discharges the whey of cumulative volume 35% after slowly stirring after 15min; Point to add 50 DEG C for 2 times, volume is the water of fresh cow milk cumulative volume 25%, stir 20min and reach 35 DEG C to curd temperature.The whey pH continuing to be stirred to discharge is 6.1, whole whey of draining.Afterwards grumeleuse is entered mould, squeezing is shaped, the demoulding is taken out.Working concentration is salt solution 12 DEG C of salt marsh 20h of 20%, and described per-cent is mass percent.Vacuum packaging afterwards, 10 DEG C, humidity 90%RH, ripe 5 weeks, obtain young Gouda cheese.
Embodiment 8 cheese starter produces Gouda cheese
1) 100kg fresh cow milk (purchased from pasture, Kingsoft) after filtration, adds 0.01% calcium chloride, stirs, and at 72 DEG C, is cooled to 30 DEG C after 15s pasteurize, and described per-cent is mass percent.Cheese starter obtained for embodiment 1 is poured in the fresh cow milk after pasteurize, makes the viable count of the cheese starter in fresh cow milk reach 10
7cfu/mL, slowly stirs 15min.Leave standstill 30 DEG C of fermentations and add 20mg/100L rennin (purchased from Danisco A/S BJ Rep Office) (wherein after pH reduces by 0.1, rennin dissolves with the sterilized water of 10 times of quality before use, stirring 2min makes it be uniformly dissolved), obtain curdled milk after 30min.
2) by step 1) obtained curd cutting becomes the cubical grumeleuse of length of side 5mm, discharges the whey of cumulative volume 35% after slowly stirring after 15min; Point to add 50 DEG C for 2 times, volume is the water of fresh cow milk cumulative volume 25%, stir 20min and reach 35 DEG C to curd temperature.The whey pH continuing to be stirred to discharge is 6.1, whole whey of draining.Afterwards grumeleuse is entered mould, squeezing is shaped, the demoulding is taken out.Working concentration is salt solution 12 DEG C of salt marsh 20h of 20%, and described per-cent is mass percent.Vacuum packaging afterwards, 10 DEG C, humidity 90%RH, ripe 15 weeks, obtain ripe Gouda cheese.
Embodiment 9 cheese starter prepares cheddar cheese-
1) with the step 1 of embodiment 7) identical.
2) by step 1) obtained curd cutting becomes the cubical grumeleuse of length of side 10mm, and place, with 1 DEG C/5min ramp to 38 DEG C, being retained to whey acidity in this temperature is 0.20%, discharging whey; Heap is made, and overturns 2 times; Stripping and slicing, adds edible sodium-chlor particulate 1.5%, and described per-cent is the mass percent accounting for cheese total mass, die-filling afterwards, vacuum packaging, 7 DEG C of maturations 12 weeks.
Embodiment 10 cheese starter prepares cheddar cheese-
1) with the step 1 of embodiment 7) identical.
2) by step 1) obtained curd cutting becomes the cubical grumeleuse of length of side 8mm, and place, with 1 DEG C/5min ramp to 40 DEG C, being retained to whey acidity in this temperature is 0.22%, discharging whey; Heap is made, and overturns 3 times; Stripping and slicing, adds edible sodium-chlor particulate 2%, and described per-cent is die-filling after being the mass percent accounting for cheese total mass, vacuum packaging, 10 DEG C of maturations 3 weeks.
Embodiment 11 cheese starter prepares cheddar cheese-
1) with the step 1 of embodiment 7) identical.
2) by step 1) obtained curd cutting becomes the cubical grumeleuse of length of side 12mm, place, with 1 DEG C/5min ramp to 38 DEG C, being retained to whey acidity in this temperature is 0.20%, discharging whey, heap is made, overturn 2 times, stripping and slicing salt adding, die-filling afterwards, vacuum packaging, 7 DEG C of maturations 12 weeks.
Embodiment 12 cheese starter prepares Pa Masen cheese
1) with the step 1 of embodiment 7) identical.
2) by step 1) obtained curd cutting becomes the cubical grumeleuse of length of side 3mm, stirs 10min.With 1 DEG C/5min ramp to 44 DEG C, insulation 10min, then continues to be warming up to 54 DEG C of insulation 5min, stirs 20min, leave standstill 20min, add a cover.Discharging whey, carries out die-filling squeezing immediately, 25 DEG C of fermentation 24h.Soak 16 days in 10 DEG C of saturated brines.Then under relative humidity 40% condition dry 1 day.16 DEG C of maturations 6 months.
Embodiment 13 cheese starter prepares Pa Masen cheese
1) with the step 1 of embodiment 7) identical.
2) by step 1) obtained curd cutting becomes the cubical grumeleuse of length of side 4mm, stirs 15min.With 2 DEG C/5min ramp to 44 DEG C, insulation 20min, then continues to be warming up to 54 DEG C of insulation 10min, stirs 10min, leave standstill 20min, add a cover.Discharging whey, carries out die-filling squeezing immediately, 25 DEG C of fermentation 24h.Soak 10 days in 16 DEG C of saturated brines.Then under relative humidity 65% condition dry 2 days.10 DEG C of ripe June.
Comparative example 1
The preparation of A, cheese starter
(1) identical with embodiment 1 step (1).
(2) culture of step (1) gained is obtained Lactococcus lactis subsp.lactis BD401, Lactococcus lactis subsp.lactis BD2263, Lactococcus lactis subsp.lactis BD164 and Leuconostoc mesenteroides LM79, then be the ratio mixing of 2:2:1:1 according to the viable count of described Lactococcus lactis subsp.lactis BD401, described Lactococcus lactis subsp.lactis BD2263, described Lactococcus lactis subsp.lactis BD164 and described Leuconostoc mesenteroides LM79, obtain cheese starter.
B, cheese starter produce Gouda cheese
Pour in the fresh cow milk after pasteurize by the cheese starter that comparative example 1 steps A obtains, all the other steps are identical with embodiment 7.Obtain young Gouda cheese.
Comparative example 2
The preparation of A, cheese starter
(1) identical with embodiment 1 step (1).
(2) culture of step (1) gained is obtained Lactococcus lactis subsp.lactis BD401, Lactococcus lactis subsp.lactis BD2263, Lactococcus lactis subsp.lactis BD164 and Leuconostoc mesenteroides LM79, then be the ratio mixing of 0.5:1:3:3 according to the viable count of described Lactococcus lactis subsp.lactis BD401, described Lactococcus lactis subsp.lactis BD2263, described Lactococcus lactis subsp.lactis BD164 and described Leuconostoc mesenteroides LM79, obtain cheese starter.
B, cheese starter produce Gouda cheese
Pour in the fresh cow milk after pasteurize by the cheese starter that comparative example 2 steps A obtains, all the other steps are identical with embodiment 7.Obtain young Gouda cheese.
Comparative example 3
The preparation of A, cheese starter
(1) identical with embodiment 1 step (1).
(2) culture of step (1) gained is obtained Lactococcus lactis subsp.lactis BD401, Lactococcus lactis subsp.lactis BD164, Lactococcus lactis subsp.lactis BD3170 and Leuconostoc mesenteroides LM79, then be the ratio mixing of 1:1:1:1 according to the viable count of described Lactococcus lactis subsp.lactis BD401, described Lactococcus lactis subsp.lactis BD164, described Lactococcus lactis subsp.lactis BD3170 and described Leuconostoc mesenteroides LM79, obtain cheese starter.
B, cheese starter produce Gouda cheese
Pour in the fresh cow milk after pasteurize by the cheese starter that comparative example 3 steps A obtains, all the other steps are identical with embodiment 7.Obtain young Gouda cheese.
Comparative example 4
The preparation of A, cheese starter
(1) identical with embodiment 1 step (1).
(2) culture of step (1) gained is obtained Lactococcus lactis subsp.lactis BD164, Lactococcus lactis subsp.lactis BD2263, Lactococcus lactis subsp.lactis BD3170 and Leuconostoc mesenteroides LM79, then be the ratio mixing of 1:1:1:1 according to the viable count of described Lactococcus lactis subsp.lactis BD164, described Lactococcus lactis subsp.lactis BD2263, described Lactococcus lactis subsp.lactis BD3170 and described Leuconostoc mesenteroides LM79, obtain cheese starter.
B, cheese starter produce Gouda cheese
Pour in the fresh cow milk after pasteurize by the cheese starter that comparative example 4 steps A obtains, all the other steps are identical with embodiment 7.Obtain young Gouda cheese.
The physical and chemical composition analysis of effect example 1 cheese
a)、pH
The cheese obtained by 10g embodiment 7 is got respectively at cheese ripening 1d, 7d, 14d, 21d, 28d, 35d and 42d, add 10mL water, homogenate (6000r/min) 5min, use pH meter (purchased from METTLER company of Switzerland) to measure the pH value of mixture, result is as shown in table 1.
The pH change during maturing of table 1 cheese
| 1st day | 7th day | 14th day | 21st day | 28th day | 35th day | 42nd day | |
| pH | 5.40 | 5.22 | 5.02 | 4.87 | 4.77 | 4.81 | 4.85 |
Table 1 illustrates, within 1 ~ 4 week ripening stage, the pH value of cheese steadily declines from 5.4 of 1d, drops to minimum, slowly rise subsequently when the 4th week (28d).This is due in Cheese during Ripening, and the milk-acid bacteria lactic acid produced that reduces lactose is consumed gradually, and the pH value of cheese presents slow ascendant trend, arrives after certain level and tends to be steady.
B), moisture
The moisture content in cheese is measured by moisture content tester MB 45 (purchased from Ohaus instrument (Shanghai) Co., Ltd.).The cheese 3g obtained by embodiment 7 of precise 42d in weighing disk, chopping is placed in sample disc, fasten capping, CHEESE pattern is set in the mode option of sample, after screen display sample initial weight, expression can run, and press Start/Stop (start/stop key), moisture content tester starts drying and measuring process.After operation completes, repeat by Display display key, screen display result record data, result is as shown in table 2.Wherein, control group for use throw type leaven CHOOZIT RM 32 (purchased from red Knicks additive (Shanghai) Co., Ltd.), according to embodiment 7 step obtained by cheese.
C), salinity
Cheese samples is ground in mortar, accurately take cheese obtained by 5g embodiment 7 in separating funnel, with the salinity in hot water repetitive scrubbing cheese, repetitive scrubbing 7 times, add hot water 25mL at every turn, washings is collected in the volumetric flask of 250mL and also uses distilled water constant volume, get 100mL washings and be placed in 250mL Erlenmeyer flask, using potassiumchromate (1mL) as colored indicator, with Silver Nitrate (0.1N) solution be titrated to become brick-red till, record consume volume V, repeat 3 times, average, result is as shown in table 2.
V-AgNO
3consumption (mL)
The weight (g) of W-cheese samples
V
1the total amount (mL) of-washings constant volume
V
2the washing lotion amount (mL) that-titration uses
D), fat
Accurately take the cheese obtained by embodiment 7 that 1.00g grinds, add 10mL 60 DEG C of warm water, be placed in 60 DEG C of water-baths and make it dissolve, add 1.25mL ammoniacal liquor, fully after mixing, keep 5min in 60 DEG C of water-baths, jolting 2min, adds 10mL ethanol, fully shakes up, be placed in after cold water cools fast and add 25mL ether, jolting 0.5min adds 25mL sherwood oil, jolting 0.5min, leaves standstill 30in, when upper liquid is clarified, read ether layer volume.Ether layer be discarded in the beaker of constant weight, be placed in boiling water bath and volatilize ether material, weigh in 100 DEG C of baking ovens after dry 1h, weigh after being again placed in the dry 40min of baking oven, front and back twice are of poor quality must not more than 1mg.Calculate by following formula, result is as shown in table 2.
Lipid content in X-sample, unit %
M
0-beaker quality, unit g
M
1-beaker adds the quality of fat, unit g
M-sample quality, unit g
The volume of V-reading ether layer, Unit/mL
The physical and chemical composition measurement result of table 2 cheese
| Cheese is numbered | Moisture (%, w/w) | FDM(％,w/w) | Salts contg (%, w/w) |
| Control group | 38.13±0.01 | 38.13±0.02 | 1.71±0.02 |
| Embodiment 7 | 39.45±0.03 | 35.29±0.01 | 1.70±0.03 |
| Comparative example 3 | 38.23±0.03 | 36.69±0.03 | 1.72±0.01 |
| Comparative example 4 | 37.25±0.02 | 37.25±0.01 | 1.69±0.01 |
E), the content of protein and hydrolysis degree thereof
Determination of total nitrogen content (TN): Kjeldahl determination, with reference to GB5009.5-2010, the reduction factor of protein is 6.38.
At the 3rd week (21d), the 6th week (42d), the cheese obtained by embodiment 7 is sampled respectively, measure pH4.6 soluble nitrogen content (pH 4.6 soluble nitrogen in the cheese obtained by embodiment 7, i.e. pH 4.6SN), 12%TCA soluble nitrogen content (12%TCA soluble nitrogen, i.e. 12%TCA-SN; Also known as non-protein nitrogen(NPN), non protein nitrogen, i.e. NPN) and the changing conditions of 5% phospho-wolframic acid soluble nitrogen (TCA-SN).
Concrete operations are as follows:
1) pH4.6 soluble nitrogen (pH4.6 SN) measures: the cheese got obtained by 10g embodiment 7 adds 40mL deionized water, carries out homogenate with high speed stamp mill, and incubation 1h under 55 DEG C of conditions also adjusts pH to be 4.6.Then at 3000g, centrifugal 30min at 4 DEG C, go upper-layer fat, collect centrifugation in centrifuge tube, preserve for-20 DEG C and be used for SDS-PAGE analysis, after supernatant liquid filtering (middling speed quantitative paper), re-use What man#42 filter paper (purchased from Whatman company) and filter, get 10mL filtrate carrying out and determine nitrogen.
2) 12% trichoroacetic acid(TCA) soluble nitrogen (TCA-SN) measures: the step 1 of 24% solution of trichloroacetic acid 25mL and 25mL pH4.6) filtrate of gained, room temperature leaves standstill 1h, again by Whatman#42 filter paper filtering, get 25mL filtrate carrying out and determine nitrogen.
3) 5% phospho-wolframic acid soluble nitrogen (PTA-SN) measures: by 5mL pH 4.6 step 1) filtrate of gained, 3.5mL 3.95M sulphuric acid soln and 1.5mL 33.3% Salkowski's solution mix, spend the night at being placed in 4 DEG C, by Whatman#42 filter paper filtering, get 10mL filtrate carrying out and determine nitrogen.
Result is as shown in table 3, wherein, control group for use throw type leaven CHOOZIT RM 32 (purchased from red Knicks additive (Shanghai) Co., Ltd.), according to embodiment 7 step obtained by cheese.
The change of proteolysis degree in table 3 Cheese during Ripening
Wherein, a, b, c and d represent and there is significant difference (p≤0.05).
Table 3 illustrates, in Cheese during Ripening, SN presents the trend rised appreciably, and this illustrates the carrying out along with maturation time, and the macromolecular polypeptides in cheese there occurs strong degraded; In Cheese during Ripening, TCA-SN presents the trend rised appreciably, and during the 3rd week ripening stage, all comparatively control group cheese is high, and the 6th week time, is tending towards close, there was no significant difference (p > 0.05); In Cheese during Ripening, PTA-SN presents the trend rised appreciably, this illustrates the carrying out along with maturation time, polypeptide in cheese etc. are degraded to amino acid gradually, and contrast control group cheese, when the 3rd and 6 weeks, its PTA-SN/TN is higher, and both also exist significant difference (p≤0.05).And comparative example 3,4 on many index all lower than control sample, particularly six weeks ripe situations are poor, and proteolysis is slow, and the flavour substances of generation is not enough.
F), water-insoluble protein SDS-PAGE
Get the step e depositing in-20 DEG C) pH 4.6 insoluble protein (0.2g) that obtains is dissolved in the solvent soln of the cheese obtained by 10mL embodiment 7 that (namely concentrated hydrochloric acid 0.4mL, mercaptoethanol 0.7mL, Tutofusin tris 0.75g, urea 49g and tetrabromophenol sulfonphthalein 0.15g are dissolved in 100mL ultrapure water, cheese obtained by embodiment 7 is dissolved in above-mentioned solution), 50 DEG C of incubation 40min, obtain incubation liquid.Then get the incubation liquid described in 1mL, the centrifugal 5min of 10000r/min, get 5 μ L supernatant liquors as sample solution.The concentration of separation gel and concentrated glue is respectively 15% and 5%, specifically in table 4.Voltage conditions is first set to 120V, is gone to bottom glue by rear voltage-regulation 100V until blue bands is gone to bottom concentrated glue to blue bands again.With coomassie brilliant blue R_250 (purchased from traditional Chinese medicines reagent), dye 2h, and then soak with destainer in shaking table, decolouring is spent the night to clear background, and result as shown in Figure 1.Wherein, control group for use throw type leaven CHOOZIT RM 32 (purchased from red Knicks additive (Shanghai) Co., Ltd.), according to embodiment 7 step obtained by cheese.
Table 4 separation gel and concentrated glue component list
As shown in Figure 1, in Cheese during Ripening, the various degradation productions of albumen produce gradually and accumulate.When cheese ripening the 6th week (42d), embodiment 7 and the electrophoresis band of the α-CN of control group are without obvious difference, and embodiment 7 is close with the β-CN electrophoresis band color of control group.
The volatile flavor substance analysis of effect example 2 cheese
Solid-phase microextraction SPME condition: the cheese 5g obtained by Example 7,30 DEG C of water bath with thermostatic control 10min, use PDMS solid phase micro-extracting head (the purchased from American Supelco company) absorption of 100um, adsorption time is 30min.
Temperature programming condition setting: keep 5min using 60 DEG C as initial temperature after sample introduction, is warming up to 120 DEG C with 5 DEG C/min and keeps 5min, and be warming up to 240 DEG C with 8 DEG C/min and keep 10min, operation total time is 47min; Chromatographic column model DB-5-MS, injector temperature 250 DEG C, ion source EI, level Four bar temperature 150 DEG C, transmission line temperature 280 DEG C, carrier gas is He, sample introduction model selection Splitless injecting samples, flow velocity 1mL/min, ionization mode EI 70eV, mass scan range 35 ~ 350m/z, result is as shown in table 5, and in table 5, the unit of numerical value is abundance.Wherein, control group for use throw type leaven CHOOZIT RM 32 (purchased from red Knicks additive (Shanghai) Co., Ltd.), according to embodiment 7 step obtained by cheese.
The measurement result of table 5 cheese main volatile flavour substances
As shown in Table 5, ketone, acid and ester compound are the main sources of cheesy flavour, wherein have 7 kinds to the compound that cheese typical flavor component contributions is maximum, i.e. hexane, 2 pentanone, ethyl butyrate, propyl carbinol, heptanone, oxaloacetic acid and butyric acid.The taste threshold of letones is lower, and has relatively high abundance in the cheese obtained by embodiment 7, larger to the local flavor integral contribution of cheese.Wherein, 3-hydroxy-2-butanone also imparts milk fragrance and the fatty taste of Gouda cheese uniqueness, and its content is in the present embodiment also higher than control group.
The sensory evaluation of effect example 3 cheese
This subjective appreciation personnel comprise the personnel that 12 are engaged in food research, are all familiar with subjective appreciation precaution and standards of grading.Total score is 50 points, and mark evaluation is in accordance with table 6.Each assessment officer independently evaluates, and use clear water to gargle during each replacing sample, result is as shown in table 7.Wherein, control group for use throw type leaven CHOOZIT RM 32 (purchased from red Knicks additive (Shanghai) Co., Ltd.), according to embodiment 7 step obtained by cheese.
Table 6 cheese subjective appreciation method
Table 7 cheese results of sensory evaluation
The result of table 7 illustrates, in the overall score growing smell and quality, the Gouda cheese (control group) that the cheese of effect example 7 gained makes than commercial fermentation agent is all slightly better.Therefore, the cheese of effect example 7 gained has good quality and better grows smell.
The quality of effect example 4 cheese
The cheese of cheese samples obtained by the embodiment 7 of ripe 60d, adopts TA-Hdi type texture analyser (purchased from Stable Micro Systems company of Britain).With sampler, cheese is cut, form diameter and be highly respectively the right cylinder fritter of 3cm, 1.5cm, being placed in lock box, in 4 DEG C of refrigerators, preserving 4h.
Location parameter: probe fall off rate 1mm/s before measuring; Test rate 1mm/s; Probe backhaul speed 1mm/s after test; Depression distance 6mm; Lower compressive strain: 50%; Trigger force: 5g; Probe type P/5.Often organize mensuration 15 times.The functional performance data (hardness, elasticity, coherency, tackyness and chewiness etc.) of cheese use texture analyser (TA-Hdi type) to carry data processing software Exponent5.0 process, and result is as shown in table 8.Wherein, control group for use throw type leaven CHOOZIT RM 32 (purchased from red Knicks additive (Shanghai) Co., Ltd.), according to embodiment 7 step obtained by cheese.
Table 8 cheese texture analysis result
Table 8 illustrates, the cheese of embodiment 7 gained in the indexs such as hardness, elasticity, chewiness and tackyness with control group cheese without significant difference, only hardness ratio contrast cheese is slightly high, illustrate that the cheese (i.e. the cheese of embodiment 7 gained) made by the cheese starter described in embodiment 1 meets the requirement of Gouda cheese in matter structure, there are the potentiality being developed to cheese starter.
Should be understood that those skilled in the art can make various changes or modifications the present invention after having read foregoing of the present invention, these equivalent form of values fall within the application's appended claims limited range equally.
Claims (10)
1. a cheese starter, it is characterized in that, it comprises Lactococcus lactis subsp.lactis (Lactococcus lactissubsp.lactis) CGMCC No.10752, Lactococcus lactis subsp.lactis (Lactococcus lactis subsp.lactis) CGMCC No.10749, Lactococcus lactis subsp.lactis (Lactococcus lactis subsp.lactis) CGMCC No.10751 and Leuconostoc mesenteroides (Leuconostoc mesenteroides) CGMCC No.10750, described Lactococcus lactis subsp.lactis CGMCC No.10752, described Lactococcus lactis subsp.lactis CGMCC No.10749, the viable count ratio of described Lactococcus lactis subsp.lactis CGMCC No.10748 and described Leuconostoc mesenteroides CGMCC No.10750 is (0.5 ~ 1): (1 ~ 1.5): (1 ~ 2): (1 ~ 2).
2. cheese starter as claimed in claim 1, it is characterized in that, the viable count ratio of described Lactococcus lactis subsp.lactis CGMCC No.10752, described Lactococcus lactis subsp.lactis CGMCC No.10749, described Lactococcus lactis subsp.lactis CGMCC No.10751 and described Leuconostoc mesenteroides CGMCC No.10750 is 1:1:1:1.
3. cheese starter as claimed in claim 1, it is characterized in that, it also comprises auxiliary material.
4. a preparation method for cheese starter, is characterized in that, it comprises the following steps:
(1) Lactococcus lactis subsp.lactis CGMCC No.10752, Lactococcus lactis subsp.lactis CGMCC No.10749, Lactococcus lactis subsp.lactis CGMCC No.10751 and Leuconostoc mesenteroides CGMCC No.10750 are inoculated in substratum, obtain culture;
(2) from the culture of step (1) gained, Lactococcus lactis subsp.lactis CGMCC No.10752 is obtained, Lactococcus lactis subsp.lactis CGMCC No.10749, Lactococcus lactis subsp.lactis CGMCC No.10748 and Leuconostoc mesenteroides CGMCC No.10750, according to Lactococcus lactis subsp.lactis CGMCC No.10752, Lactococcus lactis subsp.lactis CGMCC No.10749, the viable count of Lactococcus lactis subsp.lactis CGMCC No.10751 and Leuconostoc mesenteroides CGMCC No.10750 is (0.5 ~ 1): (1 ~ 1.5): (1 ~ 2): the ratio mixing of (1 ~ 2).
5. preparation method as claimed in claim 4, is characterized in that, comprise the step of activation before the inoculation described in step (1); The algebraically of described activation was 2 ~ 3 generations; The substratum of described activation is degreasing milk medium; The temperature of described activation is 28 ~ 32 DEG C; And/or the time of described activation is 16 ~ 24 hours.
6. preparation method as claimed in claim 5, it is characterized in that, the substratum of step (1) described activation is 10% degreasing milk medium, and described per-cent is mass percent.
7. preparation method as claimed in claim 4, it is characterized in that, the substratum described in step (1) is degreasing milk medium; The temperature of step (1) described cultivation is 28 ~ 32 DEG C; The time of step (1) described cultivation is 16 ~ 24 hours; And/or the number of times of step (1) described cultivation is 2 ~ 3 times.
8. preparation method as claimed in claim 4, it is characterized in that, in step (2), be the ratio mixing of 1:1:1:1 according to the viable count of Lactococcus lactis subsp.lactis CGMCC No.10752, Lactococcus lactis subsp.lactis CGMCC No.10749, Lactococcus lactis subsp.lactis CGMCC No.10751 and Leuconostoc mesenteroides CGMCC No.10750.
9. preparation method as claimed in claim 4, is characterized in that, further comprising the steps of after step (2) completes: to add auxiliary material, mixing.
10. the cheese starter as described in any one of claims 1 to 3 is preparing the application in cheese.
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