CN103141723A - Preparation method of lactobacillus casei soybean milk direct vat set starter - Google Patents
Preparation method of lactobacillus casei soybean milk direct vat set starter Download PDFInfo
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- CN103141723A CN103141723A CN2013100793371A CN201310079337A CN103141723A CN 103141723 A CN103141723 A CN 103141723A CN 2013100793371 A CN2013100793371 A CN 2013100793371A CN 201310079337 A CN201310079337 A CN 201310079337A CN 103141723 A CN103141723 A CN 103141723A
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Abstract
The invention discloses a preparation method of lactobacillus casei soybean milk direct vat set starter. Lactobacillus casei is subjected to high-density culture by a low-price enrichment medium, centrifugal concentration and separation, mixed dissolution with high-efficiency composite protective agent and lyophilization to prepare the lactobacillus casei soybean milk direct vat set starter. According to the preparation method, the viable organism of the prepared starter is over 1*10<11>cfu/g; after the starter is sealed under normal pressure and preserved at 4 DEG C for 1 year, the content of the viable organism is still over 10<10>cfu/g, and a higher fermentation activity is stilled maintained; and soybean milk is fermented at 37 DEG C with the inoculum size being lower than one ten-thousandth, and the milk can be cured after 4 to 5 hours, wherein the soybean yoghurt product fermented by the starter has the acidity being between 70 oT and 80oT, the pH value being about 4.6, and good sense and flavor. The starter can be used in the fermented dairy products industry and vegetable protein processing industry.
Description
Technical field
The present invention relates to a kind of preparation method of Lactobacillus casei soybean yogurt throw type leaven, belong to fermented dairy product and vegetable protein processing technique field.
Background technology
Probio means that with people and animal and plant body keep symbiosis the host is had the general name of a large quasi-microorganism of health promotion effect.At present, internationally recognized and the typical probiotic lactobacillus human body symbiosis mainly comprise: the first generation probiotic lactobacillus take Bifidobacterium and lactobacillus acidophilus as representative and take Lactobacillus casei and Lactobacillus rhamnosus as the novel probiotic lactobacillus of the second generation of representative.Than first generation probiotic lactobacillus, the novel probiotic lactobacillus of the second generation is oxytolerant, stomach juice-resistant, bile tolerance more; Fermenting property is better, viability is higher.
At present, the Lactobacillus casei fermented product is mainly the cow's milk product, and its fermented vegetable protein yogurt product still is in the research starting stage at home and abroad, and lacks the good special bacteria of production performance.
China has natural soybean resource advantage, and soybean is not only at the various functional components of nutrition and health care with promote can compare favourably with cow's milk aspect the growth factor of probio breeding, and does not contain cholesterol and lactose.Therefore, the beneficial characteristics of developing comprehensive probio and soybean, integrate the novel probio of the second generation (as Lactobacillus casei) the fermentation soybean milk product of sense organ local flavor and multiple nutrients health care, be developing direction and the new growth point of current fermented dairy product industry and soybean processing industry, will lead market and consumer to the new trend of healthy food demand.
The preparation of leavening is key link and the top priority of developing the novel fermentation dairy products.Tradition cultivation type leavening exists viable bacteria content low by (10
7～10
8Cfu/mL), inoculum concentration large (2%～5%), preservation term are short (4 ℃, 1～2d), production process is many, labour intensity is large, cultivation cycle is long, bacterial classification is very easily degenerated and the drawback such as pollution, and this will directly cause, and the fermented dairy product product quality is unstable and production efficiency is low.Thereby the novel fermentation agent---efficient concentration type throw type leaven arises at the historic moment.Efficient concentration type throw type leaven be lactic acid bacteria through deep liquid High Density Cultivation, concentrating and separating, miscible with the protective agent medium, again through freezing or freeze drying, aseptic packaging, make direct putting type cold storage leavening or direct putting type freeze dried fermenting preparation.Has viable bacteria content high by (10
10～10
12Cfu/g), long (4 ℃ of the freeze dried fermenting preparations of preservation term, 1 year), the more traditional cultivation type leavening of inoculum concentration reduces by 1000～10000 times, omitted the preparation section of leavening, investment and the space of having reduced the bacterial classification workshop, the degeneration that has prevented bacterial classification and pollution in fermented dairy product production, and the labor productivity of fermented dairy product industry and product quality are improved greatly.Direct putting type cold storage leavening is kept freezing expense higher in application, transportation is inconvenient, and is excessively strong to the dependence at leavening center.And the direct putting type freeze dried fermenting preparation, preservation and convenient transportation, mechanization degree is high, is convenient to industrialized mass production, is the main development direction of throw type leaven from now on.
The principal element that affects efficient concentration type direct putting type freeze dried fermenting preparation cell survival rate and storage-stable has: strain properties, culture medium, condition of culture and harvest time, initiator cell concentration, freeze drying protectant and pH value thereof, chilling rate, dry run, residual moisture, packaged form, preservation temperature etc.Wherein, optimizing the cheap enriched medium of screening and efficient freeze drying protectant is the key technology of development direct putting type freeze dried fermenting preparation.Studies show that: lactic acid bacteria is very harsh to nutritional requirement, the lactic acid bacteria culture medium (as MRS, M17 etc.) that the laboratory is commonly used, and complicated component is made loaded down with trivial details, expensive.At present, although degreasing milk medium and whey medium economy commonly used are easy to get, cheap, degreasing milk medium viscosity is large, is difficult for the concentrating and separating thalline; And the thermal denaturation of lactalbumin easily occurs in whey medium in the heat sterilization process and produce precipitation.Therefore, according to the china natural resources advantage, actively seek wide material sources, the cheap raw material that economy is easy to get is optimized the enriched medium that screening is fit to the lactic acid bacteria raised growth, is one of key technology that realizes China's large-scale industrial production direct putting type freeze dried fermenting preparation.Freeze drying protectant is the most complicated, the key factor of difficult selection, and it not only affects the cell survival rate of lactic acid bacteria in freeze-drying process, also affects the cell stability between preservation term, and its formula and preparation method thereof belongs to trade secret.Therefore, optimize the efficient freeze drying protectant of screening be develop the direct putting type freeze dried fermenting preparation key technology two.
As fully visible, actively develop the efficient concentration type direct putting type probio soy acid milk fermentation agent with autonomous innovation of alternative imported product, for the quality that improves the fermentation soybean milk product and functional, promote the technological progress of China's efficient concentration type direct putting type probiotics leaven industrialization, promote the development of China's fermentation soybean dairy products and vegetable protein secondary industry, significant.
Application number be 200410018684.4 Chinese disclosure of the invention " preparation method of probiotic lactobacillus leavening and the application in fresh cheese thereof ", through High Density Cultivation with a strain Lactobacillus casei, centrifugal concentrating, carry out freeze drying after miscible with protective agent and obtain freeze dried fermenting preparation, but its modified MRS culture medium complicated component used, make loaded down with trivial detailsly, cost is higher; And clearly whether this bacterium is not applicable to the fermentation soybean yogurt yet.Application number has been 200810175856.7 Chinese disclosure of the invention " Lactobacillus casei Zhang high-density cultivation method, use them to prepare the method for freeze-dried vaccine powder and resulting freeze-dried vaccine powder and uses thereof ", the freeze-dried vaccine powder that has prepared Lactobacillus casei Zhang, but its high-density culture medium complicated component used, make loaded down with trivial detailsly, cost is higher; Freeze-drying time is 36h, and efficient awaits further raising, and does not relate to this bacterium and whether also be applicable to the fermentation soybean yogurt.
Summary of the invention
The preparation method who the purpose of this invention is to provide a kind of Lactobacillus casei soybean yogurt throw type leaven is to realize the industrial-scale production of Lactobacillus casei throw type leaven.
The applicant has carried out collection, the separation screening work of a large amount of novel probios in earlier stage, growth to novel probio in Soybean Milk, fermentation character etc. study in great detail, discovery comes from the Lactobacillus casei AS1.62 of Institute of Microorganism, Academia Sinica, have good growth and fermentation character in the Soybean Milk culture medium, cultivate 12 hours viable counts and can reach 10
9Cfu/mL, and it has been carried out follow-up soybean yogurt throw type leaven preparation method's research.For the characteristics of lactic acid bacteria to the nutritional requirement harshness, by lot of experiments, select that colloidal state cheap and easy to get---jerusalem artichoke is raw material, studied the preparation technology of jerusalem artichoke juice basal medium; And take Lactobacillus casei AS1.62 as test strain, studied at the jerusalem artichoke juice basal medium and added single trophic factors to the impact of Lactobacillus casei cell yield, further utilized optimization of orthogonal test to filter out the compound enriched medium of jerusalem artichoke juice of Lactobacillus casei; By the impact of the factors such as research inoculum concentration, cultivation temperature, matrix Initial pH, training method on the Lactobacillus casei cell yield, further utilize optimization of orthogonal test to determine the process conditions that Lactobacillus casei propagation is cultivated; Study centrifugal force, centrifugation time to the impact of cell centrifugation loss late and survival rate, determined the suitableeest centrifuging process condition of the highest viable bacteria recovery rate; Meanwhile, on the lot of experiments basis, studied different composite protectants to the impact of Lactobacillus casei freeze-drying survival rate, optimized and filter out anti-cryodesiccated complex protection agent prescription; The Lactobacillus casei freeze dried powder that the anti-cryodesiccated complex protection agent prescription that optimization is filtered out is made by the cell accelerated test, further filters out not only anti-freeze drying but also long keeping good complex protection agent prescription.Add Lactobacillus casei bacteria suspension chilling rate impact on cell survival rate in refrigerating process of composite protectant by research, determined the freeze drying process of reasonable; By research Lactobacillus casei freeze dried fermenting preparation seal the lower packing of mode (normal pressure is sealed up for safekeeping, vacuum preservation) up for safekeeping in difference, in the viable count of the lower storage of different temperatures (18 ℃, 4 ℃) and the variation of fermentative activity, determined storage conditions and the preservation term of freeze dried fermenting preparation.
Concrete, the preparation method of a kind of Lactobacillus casei soybean yogurt throw type leaven provided by the invention comprises the following steps:
(1) propagation of Lactobacillus casei is cultivated
1. the preparation of the cheap enriched medium of Lactobacillus casei
A. the preparation of Lactobacillus casei jerusalem artichoke juice basal medium
Raw material: jerusalem artichoke, distilled water;
Preparation technology: (a) jerusalem artichoke is cleaned, weighs; (b) jerusalem artichoke is that 1:1 mixes with water by mass volume ratio; (c) heat the enzyme that goes out: be heated to 100 ℃, boil 5min; (d) adding water smashs to pieces: add appropriate 80 ℃ of hot water, smash jerusalem artichoke and water to pieces pulp in bruisher; (e) filter: with 100 order filter cloth press filtrations; (f) filter residue adds the hydraulic pressure press for extracting juice: filter residue is added appropriate 80 ℃ of hot water press filtrations; (g) filtered juice mixing, constant volume: the filtered juice of twice filtration is mixed, be settled to the 1kg jerusalem artichoke with distilled water and produce 1L juice (being defined as 100% jerusalem artichoke juice); (h) allotment: it is 10% that the jerusalem artichoke juice heating is concentrated into sugar content, regulates pH to 7.0 with NaOH solution, makes Lactobacillus casei jerusalem artichoke juice basal medium, standby;
B. the preparation of the compound enriched medium of Lactobacillus casei
Raw material and proportioning: Lactobacillus casei jerusalem artichoke juice basal medium 100mL, yeast extract 0.5～0.8g, peptone 0.3～0.6g, soy peptone 1.2～1.6g, beef extract 0.8～1.2g, glycerine 0.1mL;
Preparation technology: yeast extract, peptone, soy peptone, beef extract, glycerine are added in Lactobacillus casei jerusalem artichoke juice basal medium, regulate pH value to 6.5～7.0,121 ℃ of 15min sterilizations, cooling, standby with NaOH solution;
2. Lactobacillus casei propagation is cultivated
Lactobacillus casei is through MRS(Man Rogosa Sharpe) after fluid nutrient medium activation, with 0.1%～0.5%(approximately 1 * 10
6Cfu/mL～5 * 10
6Cfu/mL) inoculum concentration access pH value is that in 6.5～7.0 the compound enriched medium of jerusalem artichoke juice, 35～37 ℃ of cultivation 16～18h are to logarithm latter stage or stable initial stage under aerobic conditions, and viable count can reach 4～5 * 10
9Cfu/mL, the high-concentration culturing thing of acquisition Lactobacillus casei;
(2) concentrating and separating of Lactobacillus casei cell
The Lactobacillus casei high-concentration culturing thing centrifugal 10～20min of 5000～7500r/min(2550g～5738g) at normal temperatures carries out cell concentration and separates, and centrifugal rear abandoning supernatant obtains the viable bacteria recovery rate higher than 95% Lactobacillus casei concentrating cells;
(3) add composite protectant in the Lactobacillus casei concentrating cells
Complex protection agent prescription: distilled water 100mL, cycloheptaamylose 10g, sucrose 6～7g, glycerine 0.1～0.2g, dusty yeast 1.0～1.2g, vitamin E 0.3～0.6g;
Composite protectant preparation technology: 1. add sucrose, glycerine, dusty yeast in distilled water, the heating for dissolving mixing is cooled to room temperature; 2. regulate pH value to 6.8～7.0 with NaOH solution; 3. 115 ℃ of sterilization 10min, be cooled to room temperature; 4. add under aseptic condition through 105 ℃, 60min hot air sterilization and cooled cycloheptaamylose and the vitamin E after degerming after filtration, mixing;
Add composite protectant technique in the Lactobacillus casei concentrating cells: add the composite protectant of bacterial strain high-concentration culturing object long-pending 20%～30% in the Lactobacillus casei concentrating cells, make bacteria suspension, mixing gets final product;
(4) freeze-dry process of Lactobacillus casei throw type leaven
The Lactobacillus casei bacteria suspension that has added composite protectant is sub-packed in freeze-drying dish or bottle thickness 0.75cm ,-20 ℃ of freezing 12h; In vacuum 3～10Pa, condenser temperature-55～-45 ℃ lower vacuum freeze drying 11～13h, make Lactobacillus casei freeze dried fermenting preparation finished product in freeze drier, the viable count of freeze dried fermenting preparation can reach 1 * 10
11More than cfu/g, water content is 2～3%.
The beneficial effect that the present invention obtains is as follows:
1. the efficient concentration type direct putting type Lactobacillus casei leavening of the present invention preparation, its fermenting property and technical indicator can compare favourably with internationally renowned brand business leavening, and (viable bacteria content can reach 1 * 10
11More than cfu/g; Normal pressure seals up for safekeeping, 4 ℃ of lower preservations 1 year, and its viable bacteria content is still 10
10More than cfu/g, still keep higher fermentative activity), and cost reduces more than 30% than import business leavening.
2. the Lactobacillus casei freeze dried fermenting preparation of the present invention preparation, can be used as the throw type leaven of producing the soybean yogurt uses, the more traditional cultivation type leavening of inoculum concentration reduces by 10000～100000 times, omitted the preparation section of leavening in industrial production, investment and the space in bacterial classification workshop have been reduced, prevent degeneration and the pollution of bacterial classification, can improve labor productivity and the product quality of fermented dairy product industry and vegetable protein secondary industry.
3. the Lactobacillus casei freeze dried fermenting preparation of the present invention's preparation, carry out 37 ℃ of bean yoghourt fermentations with ten thousand/following inoculum concentration, and 4～5h gets final product curdled milk, the bean yoghourt goods of its fermentation, and acidity reaches 70～80
oT, the pH value is 4.6 left and right, the sense organ raciness.Can be used for fermented dairy product industry and vegetable protein secondary industry.
Description of drawings
Fig. 1 be after the Lactobacillus casei soybean yogurt throw type leaven normal pressures of embodiment 1,2,3 preparations are sealed up for safekeeping under 4 ℃ and-18 ℃ of conditions the storage viable count of 1 year change.
The specific embodiment
Following examples are used for explanation the present invention.
The preparation method of 1 one kinds of Lactobacillus casei soybean yogurt throw type leavens of embodiment, realize by following steps:
(1) propagation of Lactobacillus casei is cultivated
1. the preparation of the cheap enriched medium of Lactobacillus casei
A. the preparation of Lactobacillus casei jerusalem artichoke juice basal medium
Raw material proportioning: jerusalem artichoke, distilled water;
Preparation technology: (a) jerusalem artichoke is cleaned, weighs; (b) mass volume ratio of material water is 1:1; (c) heat the enzyme that goes out: be heated to 100 ℃, boil 5min; (d) adding water smashs to pieces: add appropriate 80 ℃ of hot water, smash jerusalem artichoke and water to pieces pulp in bruisher; (e) filter: with 100 order filter cloth press filtrations; (f) filter residue adds the hydraulic pressure press for extracting juice: filter residue is added appropriate 80 ℃ of hot water press filtrations; (g) mix the filtered juice constant volume: the filtered juice of twice filtration is mixed, be settled to the 1kg jerusalem artichoke with distilled water and produce 1L juice (being defined as 100% jerusalem artichoke juice); (h) allotment: it is 10% left and right that heating is concentrated into sugar content; Regulate pH to 7.0 left and right with NaOH solution;
B. the preparation of the compound enriched medium of Lactobacillus casei
Raw material proportioning: Lactobacillus casei jerusalem artichoke juice basal medium 100mL, yeast extract 0.5g, peptone 0.3g, soy peptone 1.6g, beef extract 1.2g, glycerine 0.1mL, pH6.5～7.0;
Preparation technology: add yeast extract 0.5g, peptone 0.3g, soy peptone 1.6g, beef extract 1.2g, glycerine 0.1mL in 100mL jerusalem artichoke juice basal medium, regulate pH value to 6.5～7.0,121 ℃ of 15min sterilizations with NaOH solution cooling standby;
2. Lactobacillus casei is bred the process conditions of cultivating
Lactobacillus casei is after MRS fluid nutrient medium activation, with 0.5%(approximately 5 * 10
6Cfu/mL) inoculum concentration access pH value is that in 6.5～7.0 the compound enriched medium of jerusalem artichoke juice, 37 ℃ of cultivation 16h are to logarithm latter stage or stable initial stage under aerobic conditions, and viable count can reach 4.3 * 10
9Cfu/mL, the high-concentration culturing thing of acquisition Lactobacillus casei;
(2) concentrating and separating of Lactobacillus casei cell
Lactobacillus casei high-concentration culturing thing adopts 7500r/min(5738g at normal temperatures) centrifugal 10min carries out cell concentration and separates, and centrifugal rear abandoning supernatant obtains the viable bacteria recovery rate and is the concentrating cells of 95.34% Lactobacillus casei;
(3) add composite protectant in the Lactobacillus casei concentrating cells
The complex protection agent prescription: distilled water 100mL, cycloheptaamylose 10g, sucrose 6g, glycerine 0.2g, dusty yeast 1.2g, vitamin E 0.3g, pH value 6.8～7.0 is filtered or the heating degerming gets final product;
The preparation technology of complex protection agent prescription: 1. add sucrose 6g, glycerine 0.2g, dusty yeast 1.2g in 100mL distilled water, the heating for dissolving mixing is cooled to room temperature; 2. regulate pH value to 6.8～7.0 with NaOH solution; 3. 115 ℃ of sterilization 10min, be cooled to room temperature; 4. adopt sterile working to add through the vitamin E 0.3g after 105 ℃ of 60min hot air sterilizations and cooled cycloheptaamylose 10g and process filtration sterilization, mixing gets final product;
Add composite protectant technique in the Lactobacillus casei concentrating cells: add the composite protectant of bacterial strain high-concentration culturing object long-pending 30% in the Lactobacillus casei concentrating cells, make the Lactobacillus casei bacteria suspension, mixing;
(4) freeze-dry process of Lactobacillus casei throw type leaven
With the protectant Lactobacillus casei bacteria suspension of interpolation packing freeze-drying dish/bottle, thickness 0.75cm ,-20 ℃ of freezing 12h; In vacuum 3～10Pa, condenser temperature-55～-45 ℃ lower vacuum freeze drying 13h, make the Lactobacillus casei freeze dried fermenting preparation in freeze drier.The viable count of freeze dried fermenting preparation can reach 1.19 * 10
11Cfu/g, water content is 2.1%.
The preparation method of 2 one kinds of Lactobacillus casei soybean yogurt throw type leavens of embodiment, the method realizes by following steps:
(1) propagation of Lactobacillus casei is cultivated
1. the preparation of the cheap enriched medium of Lactobacillus casei
A. the raw material proportioning of Lactobacillus casei jerusalem artichoke juice basal medium and preparation technology are with embodiment 1.
B. the preparation of the compound enriched medium of Lactobacillus casei
Raw material proportioning: Lactobacillus casei jerusalem artichoke juice basal medium 100mL, yeast extract 0.7g, peptone 0.5g, soy peptone 1.5g, beef extract 1.0g, pH6.5～7.0;
Preparation technology: add yeast extract 0.7g, peptone 0.5g, soy peptone 1.5g, beef extract 1.0g, glycerine 0.1mL in 100mL jerusalem artichoke juice basal medium, regulate pH value to 6.5～7.0,121 ℃ of 15min sterilizations with NaOH solution cooling standby;
2. Lactobacillus casei is bred the process conditions of cultivating
Lactobacillus casei is after MRS fluid nutrient medium activation, with 0.1%(approximately 1 * 10
6Cfu/mL) inoculum concentration access pH value is that in 6.5～7.0 the compound enriched medium of jerusalem artichoke juice, 35 ℃ of cultivation 18h are to logarithm latter stage or stable initial stage under aerobic conditions, and viable count can reach 4.85 * 10
9Cfu/mL, the high-concentration culturing thing of acquisition Lactobacillus casei;
(2) concentrating and separating of Lactobacillus casei cell
Lactobacillus casei high-concentration culturing thing adopts 5000r/min(2550g at normal temperatures) centrifugal 20min carries out cell concentration and separates, and centrifugal rear abandoning supernatant obtains the viable bacteria recovery rate and is 98.35% Lactobacillus casei concentrating cells;
(3) add composite protectant in the Lactobacillus casei concentrating cells
The complex protection agent prescription: distilled water 100mL, cycloheptaamylose 10g, sucrose 6～7g, glycerine 0.1～0.2g, dusty yeast 1.0～1.2g, vitamin E 0.3～0.6g, pH value 6.8～7.0 is filtered or the heating degerming gets final product;
The preparation technology of complex protection agent prescription: 1. add sucrose 6g, glycerine 0.1g, dusty yeast 1.0g in 100mL distilled water, the heating for dissolving mixing is cooled to room temperature; 2. regulate pH value to 6.8～7.0 with NaOH solution; 3. 115 ℃ of sterilization 10min, be cooled to room temperature; 4. adopt sterile working to add through the vitamin E 0.5g after 105 ℃ of 60min hot air sterilizations and cooled cycloheptaamylose 10g and process filtration sterilization, mixing gets final product;
Add composite protectant technique in the Lactobacillus casei concentrating cells: add the composite protectant of bacterial strain high-concentration culturing object long-pending 20% in the Lactobacillus casei concentrating cells, make bacteria suspension, mixing gets final product;
(4) freeze-dry process of Lactobacillus casei throw type leaven
With the protectant Lactobacillus casei bacteria suspension of interpolation packing freeze-drying dish/bottle, thickness 0.75cm ,-20 ℃ of freezing 12h; In vacuum 3～10Pa, condenser temperature-55～-45 ℃ lower vacuum freeze drying 11h, make the Lactobacillus casei freeze dried fermenting preparation in freeze drier.The viable count of freeze dried fermenting preparation can reach 1.43 * 10
11Cfu/g, water content is 2.7%.
The preparation method of 3 one kinds of dried leavenings of Lactobacillus casei direct putting type soy acid junket of embodiment, the method realizes by following steps:
(1) propagation of Lactobacillus casei is cultivated
1. the preparation of the cheap enriched medium of Lactobacillus casei
A. the raw material proportioning of Lactobacillus casei jerusalem artichoke juice basal medium and preparation technology are with embodiment 1.
B. the preparation of the compound enriched medium of Lactobacillus casei
Raw material proportioning: Lactobacillus casei jerusalem artichoke juice basal medium 100mL, yeast extract 0.8g, peptone 0.6g, soy peptone 1.2g, beef extract 0.8g, glycerine 0.1mL, pH6.5～7.0;
Preparation technology: add yeast extract 0.8g, peptone 0.6g, soy peptone 1.2g, beef extract 0.8g, glycerine 0.1mL in 100mL jerusalem artichoke juice basal medium, regulate pH value to 6.5～7.0,121 ℃ of 15min sterilizations with NaOH solution cooling standby;
2. Lactobacillus casei is bred the process conditions of cultivating
Lactobacillus casei is after MRS fluid nutrient medium activation, with 0.3%(approximately 1 * 10
6Cfu/mL) inoculum concentration access pH value is that in 6.5～7.0 the compound enriched medium of jerusalem artichoke juice, 37 ℃ of cultivation 17h are to logarithm latter stage or stable initial stage under aerobic conditions, and viable count can reach 4.3 * 10
9Cfu/mL, the high-concentration culturing thing of acquisition Lactobacillus casei;
(2) concentrating and separating of Lactobacillus casei cell
Lactobacillus casei high-concentration culturing thing adopts 6000r/min(3672g at normal temperatures) centrifugal 15min carries out cell concentration and separates, and centrifugal rear abandoning supernatant obtains the viable bacteria recovery rate and is 96.05% Lactobacillus casei concentrating cells;
(3) add composite protectant in the Lactobacillus casei concentrating cells
The complex protection agent prescription: distilled water 100mL, cycloheptaamylose 10g, sucrose 7g, glycerine 0.1g, dusty yeast 1.0g, vitamin E 0.6g, pH value 6.8～7.0 is filtered or the heating degerming gets final product;
The preparation technology of complex protection agent prescription: 1. add sucrose 7g, glycerine 0.1g, dusty yeast 1.0g in 100mL distilled water, the heating for dissolving mixing is cooled to room temperature; 2. regulate pH value to 6.8～7.0 with NaOH solution; 3. 115 ℃ of sterilization 10min, be cooled to room temperature; 4. adopt sterile working to add through the vitamin E 0.6g after 105 ℃ of 60min hot air sterilizations and cooled cycloheptaamylose 10g and process filtration sterilization, mixing gets final product;
Add composite protectant technique in the Lactobacillus casei concentrating cells: add the composite protectant of bacterial strain high-concentration culturing object long-pending 25% in the Lactobacillus casei concentrating cells, make the Lactobacillus casei bacteria suspension, mixing;
(4) freeze-dry process of Lactobacillus casei throw type leaven
With the protectant Lactobacillus casei bacteria suspension of interpolation packing freeze-drying dish/bottle, thickness 0.75cm ,-20 ℃ of freezing 12h; In vacuum 3～10Pa, condenser temperature-55～-45 ℃ lower vacuum freeze drying 12h, make the Lactobacillus casei freeze dried fermenting preparation in freeze drier.The viable count of freeze dried fermenting preparation can reach 1.24 * 10
11Cfu/g, water content is 2.4%.
The subordinate list explanation:
Table 1 for seal up for safekeeping at normal pressure, the Lactobacillus casei throw type leaven of-18 ℃ of storages is on the impact of soy acid milk fermentation and quality.
Claims (1)
1. the preparation method of a Lactobacillus casei soybean yogurt throw type leaven is characterized in that comprising the following steps:
(1) propagation of Lactobacillus casei is cultivated
1. the preparation of the cheap enriched medium of Lactobacillus casei
A. the preparation of Lactobacillus casei jerusalem artichoke juice basal medium
Raw material: jerusalem artichoke, distilled water;
Preparation technology: (a) jerusalem artichoke is cleaned, weighs; (b) jerusalem artichoke is that 1:1 mixes with water by mass volume ratio; (c) heat the enzyme that goes out: be heated to 100 ℃, boil 5min; (d) adding water smashs to pieces: add appropriate 80 ℃ of hot water, smash jerusalem artichoke and water to pieces pulp in bruisher; (e) filter: with 100 order filter cloth press filtrations; (f) filter residue adds the hydraulic pressure press for extracting juice: filter residue is added appropriate 80 ℃ of hot water press filtrations; (g) filtered juice mixing, constant volume: the filtered juice of twice filtration is mixed, be settled to the 1kg jerusalem artichoke with distilled water and produce 1L juice; (h) allotment: it is 10% that the jerusalem artichoke juice heating is concentrated into sugar content, regulates pH to 7.0 with NaOH solution, makes Lactobacillus casei jerusalem artichoke juice basal medium, standby;
B. the preparation of the compound enriched medium of Lactobacillus casei
Raw material and proportioning: Lactobacillus casei jerusalem artichoke juice basal medium 100mL, yeast extract 0.5～0.8g, peptone 0.3～0.6g, soy peptone 1.2～1.6g, beef extract 0.8～1.2g, glycerine 0.1mL;
Preparation technology: yeast extract, peptone, soy peptone, beef extract, glycerine are added in Lactobacillus casei jerusalem artichoke juice basal medium, regulate pH value to 6.5～7.0,121 ℃ of 15min sterilizations, cooling, standby with NaOH solution;
2. Lactobacillus casei propagation is cultivated
Lactobacillus casei is after the activation of MRS fluid nutrient medium, access the pH value in the compound enriched medium of 6.5～7.0 jerusalem artichoke juice take 0.1%～0.5% inoculum concentration, cultivate 16～18h to logarithm latter stage or stable initial stage for 35～37 ℃ under aerobic conditions, viable count reaches 4～5 * 10
9Cfu/mL, the high-concentration culturing thing of acquisition Lactobacillus casei;
(2) concentrating and separating of Lactobacillus casei cell
The Lactobacillus casei high-concentration culturing thing centrifugal 10～20min of 5000～7500r/min at normal temperatures carries out cell concentration and separates, and centrifugal rear abandoning supernatant obtains the viable bacteria recovery rate higher than 95% Lactobacillus casei concentrating cells;
(3) add composite protectant in the Lactobacillus casei concentrating cells
Complex protection agent prescription: distilled water 100mL, cycloheptaamylose 10g, sucrose 6～7g, glycerine 0.1～0.2g, dusty yeast 1.0～1.2g, vitamin E 0.3～0.6g;
Composite protectant preparation technology: 1. add sucrose, glycerine, dusty yeast in distilled water, the heating for dissolving mixing is cooled to room temperature; 2. regulate pH value to 6.8～7.0 with NaOH solution; 3. 115 ℃ of sterilization 10min, be cooled to room temperature; 4. add under aseptic condition through 105 ℃, 60min hot air sterilization and cooled cycloheptaamylose and the vitamin E after degerming after filtration, mixing;
Add composite protectant technique in the Lactobacillus casei concentrating cells: add the composite protectant of bacterial strain high-concentration culturing object long-pending 20%～30% in the Lactobacillus casei concentrating cells, make the Lactobacillus casei bacteria suspension, mixing;
(4) freeze-dry process of Lactobacillus casei throw type leaven
The Lactobacillus casei bacteria suspension that has added composite protectant is sub-packed in freeze-drying dish or bottle thickness 0.75cm ,-20 ℃ of freezing 12h; In vacuum 3～10Pa, condenser temperature-55～-45 ℃ lower vacuum freeze drying 11～13h, make Lactobacillus casei freeze dried fermenting preparation finished product in freeze drier.
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