CN108112890A - A kind of enzymatic hydrolysis and fermentation bone meal and preparation method thereof - Google Patents
A kind of enzymatic hydrolysis and fermentation bone meal and preparation method thereof Download PDFInfo
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- CN108112890A CN108112890A CN201711374754.3A CN201711374754A CN108112890A CN 108112890 A CN108112890 A CN 108112890A CN 201711374754 A CN201711374754 A CN 201711374754A CN 108112890 A CN108112890 A CN 108112890A
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- bone meal
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- 239000002374 bone meal Substances 0.000 title claims abstract description 143
- 229940036811 bone meal Drugs 0.000 title claims abstract description 143
- 238000000855 fermentation Methods 0.000 title claims abstract description 136
- 230000004151 fermentation Effects 0.000 title claims abstract description 136
- 230000007071 enzymatic hydrolysis Effects 0.000 title claims abstract description 57
- 238000006047 enzymatic hydrolysis reaction Methods 0.000 title claims abstract description 57
- 238000002360 preparation method Methods 0.000 title claims abstract description 20
- 229940088598 enzyme Drugs 0.000 claims abstract description 66
- 102000004190 Enzymes Human genes 0.000 claims abstract description 63
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- 239000004365 Protease Substances 0.000 claims abstract description 51
- 210000000988 bone and bone Anatomy 0.000 claims abstract description 50
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- 239000011669 selenium Substances 0.000 claims abstract description 36
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- 229910052711 selenium Inorganic materials 0.000 claims abstract description 30
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- 239000002994 raw material Substances 0.000 claims abstract description 19
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- 241000186672 Lactobacillus delbrueckii subsp. bulgaricus Species 0.000 claims abstract 3
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- 238000006460 hydrolysis reaction Methods 0.000 claims description 89
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- 235000015510 Cucumis melo subsp melo Nutrition 0.000 description 2
- FJJCIZWZNKZHII-UHFFFAOYSA-N [4,6-bis(cyanoamino)-1,3,5-triazin-2-yl]cyanamide Chemical compound N#CNC1=NC(NC#N)=NC(NC#N)=N1 FJJCIZWZNKZHII-UHFFFAOYSA-N 0.000 description 2
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- 206010039921 Selenium deficiency Diseases 0.000 description 1
- 238000000944 Soxhlet extraction Methods 0.000 description 1
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- 150000001413 amino acids Chemical class 0.000 description 1
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- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical compound [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 description 1
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Classifications
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
- A23L13/00—Meat products; Meat meal; Preparation or treatment thereof
- A23L13/20—Meat products; Meat meal; Preparation or treatment thereof from offal, e.g. rinds, skins, marrow, tripes, feet, ears or snouts
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
- A23L13/00—Meat products; Meat meal; Preparation or treatment thereof
- A23L13/40—Meat products; Meat meal; Preparation or treatment thereof containing additives
- A23L13/48—Addition of, or treatment with, enzymes
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
- A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
- A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
- A23L33/16—Inorganic salts, minerals or trace elements
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
- A23V2002/00—Food compositions, function of food ingredients or processes for food or foodstuffs
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- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Health & Medical Sciences (AREA)
- Nutrition Science (AREA)
- Engineering & Computer Science (AREA)
- Food Science & Technology (AREA)
- Polymers & Plastics (AREA)
- Inorganic Chemistry (AREA)
- Mycology (AREA)
- Microbiology (AREA)
- Coloring Foods And Improving Nutritive Qualities (AREA)
Abstract
The invention belongs to bone meal food technical fields, specifically disclose a kind of enzymatic hydrolysis and fermentation bone meal and preparation method thereof, the enzymatic hydrolysis and fermentation bone meal is after selenium-rich ox bone raw material is made degreasing fermentation bone meal, through neutral proteinase and papain enzymolysis, by lactobacillus bulgaricus and the fermented by mixed bacterium of streptococcus thermophilus, finally it is dried to obtain.The present invention is for the first time by neutral proteinase and the double enzyme combination enzymatic hydrolysis and fermentation bovine bone powders of papain, quality improving, mitigate bone meal granular sensation, bone meal is more smooth, it is determined that enzymolysis and the optimum condition of fermentation impart ferment local-flavor after lactobacillus-fermented to bone meal, macromolecular substances are decomposed, improve flavor and improve nutritional quality, value-added content of product is high, improves the value and utilization rate of ox bone.
Description
Technical field
The invention belongs to bone meal food technical fields, and in particular to a kind of enzymatic hydrolysis and fermentation bone meal and preparation method thereof.
Background technology
The investigation of the version Chinese sirloin market statuses in 2017 and trend analysis report of China's Industry investigation net issue are recognized
For the U.S., Brazil and China are the countries that beef consumes first three.China's beef production reaches 6,800,000 tons within 2016, in recent years,
The yield of China's beef is into the trend of cumulative year after year.As the Inner Mongol first of five pastoral area of China, end 11 the end of month in 2016, entirely
Area ox delivers 417.2 ten thousand for sale, increases by 7.1% on a year-on-year basis.71.3 ten thousand tons of beef production, increases by 8.9% on a year-on-year basis.Ox bone accounts for Niu Tichong's
10%-20%, therefore, through rough Statistics, the yield of China's ox bone in 2016 is about between ten thousand tons of 7.9-17.8.It can be seen that China
Ox bone yield is big.But since ox bone difficulty of processing is big, research and development ox bone historical time is shorter, and storage difficulty is big, and beef cattle is slaughtered
Beef cattle byproduct processing and utilization is realized the influence of the factors such as not strong by slaughterhouse enterprise, and substantial amounts of ox bone is not developed effectively
Utilize, can only add in feed as livestock calcium tonic or be processed into the poor byproduct of mouthfeel, and high added value is excellent
The ox bone product of matter is less.
The country is compared to, it is external more extensive to the research of ox bone, such as the states such as Germany, Britain, Italy and France
Utilize ox bone edible protein, bone collagen and animal gelatin；Ultramicro grinding bone meal is added to baby food, beef by the U.S.
In the food such as gruel, minced beef cutlet, butcher's meat.
Selenium element is one of needed by human 15 kind element that the United Nations health organization is established, and is that human body can not synthesize and must
Must be by the trace element of food intake, the effect of requirement very little is huge, is known as " the atom in body trace element by scientific circles
Bullet ".Its six macro biologies function is：1st, anti-oxidant, anti-aging, 2, protection, repair cell, 3, improve red blood cell oxygen carrying energy
Power, 4, improve the immunity of the human body, 5, removing toxic substances, toxin expelling, antipollution, 6, the prevention canceration World Health Organization claims：People should be as intake
Protein is the same, must take in 50-250 microgram selenium daily, is to supplement by natural organic selenium-selenium-enriched food in food
Most scientific.Due to the universal selenium deficiency of Chinese wide geographic area, it is most easily method to supplement selenium element by selenium-enriched food, institute
With in recent years, the exploitation and popularization of selenium-enriched food are always one of research hotspot of functional food.
Therefore, selenium-rich ox bone product of the research and development with high added value, improves the value and utilization rate of ox bone, can make China's meat
The profit of ox enterprise is further promoted.
The content of the invention
A kind of enzymatic hydrolysis and fermentation bone meal provided by the invention and preparation method thereof, the enzymolysis being prepared into using double enzyme enzymolysis process
Ferment bone meal, and value-added content of product is high, improves the value and utilization rate of ox bone.
The object of the present invention is to provide a kind of enzymatic hydrolysis and fermentation bone meal, the enzymatic hydrolysis and fermentation bone meal is by selenium-rich ox bone raw material system
After fermenting bone meal into degreasing, through neutral proteinase and papain enzymolysis, by lactobacillus bulgaricus and streptococcus thermophilus
Fermented by mixed bacterium, be finally dried to obtain.
The present invention also provides a kind of preparation methods of enzymatic hydrolysis and fermentation bone meal, comprise the following steps：
S1 chooses selenium-rich ox bone raw material, prepares degreasing fermentation bone meal；
S2, enzymolysis
The bone tankage solution of 10g/100ml is prepared, adjusting pH value is 6-8, rear to add in neutral proteinase and Papain
Enzyme, enzymolysis, after digesting, is placed in 90 DEG C of enzyme deactivation 20min by enzymolysis liquid, obtains enzymatic hydrolysis and fermentation bone meal solution；
Wherein, the additive amount of neutral proteinase is 10-14g/kg, papain 10-16g/kg, hydrolysis temperature 50-
55 DEG C, enzymolysis time 4-6h；
S3, fermentation
Glucose is added in into enzymatic hydrolysis and fermentation bone meal solution, adjusting pH value is 6-8, and sterilizing is then respectively adding activation
Lactobacillus bulgaricus seed liquor and the streptococcus thermophilus seed liquor of activation, carry out fermentation 24-30h, obtain fermentation bone meal solution；
Wherein, the usage ratio of enzymatic hydrolysis and fermentation bone meal solution and glucose is 100ml:5g, lactobacillus bulgaricus seed
The volumetric usage ratio of liquid and streptococcus thermophilus seed liquor is 2:1；
Fermentation bone meal solution is evaporated, seasons, enzymatic hydrolysis and fermentation bone meal is made by S4.
Preferably, the preparation method of above-mentioned enzymatic hydrolysis and fermentation bone meal, the technique for preparing degreasing fermentation bone meal are as follows：
S11, fermentation bone meal are crude
Selenium-rich ox bone raw material through degreasing, removes marrow, animal oil successively, air-dries, smashs to pieces, sieve after grinds, obtain crude
Ferment bone meal；
S12, bone meal degreasing of fermenting
Crude fermentation bone meal is added in the ethyl alcohol that volumetric concentration is 95%, solid-liquid ratio 1g:15ml, degreasing, most
After air-dry, obtain degreasing fermentation bone meal.
Preferably, the preparation method of above-mentioned enzymatic hydrolysis and fermentation bone meal, S11's concretely comprises the following steps：Selenium-rich ox bone raw material passes through successively
Boiling water normal pressure infusion 120min degreasings, 0.1MPa autoclaving 40min remove marrow, animal oil, and 60-70 DEG C of air-dried 10h is smashed to pieces, mill
Cheng Fenhou crosses 100 mesh sieves, obtains crude fermentation bone meal.
Preferably, the preparation method of above-mentioned enzymatic hydrolysis and fermentation bone meal, in S12, time of degreasing is 9h, temperature is 45 DEG C.
Preferably, the preparation method of above-mentioned enzymatic hydrolysis and fermentation bone meal, in S2, the vigor of neutral proteinase is 11036U/g, wood
The vigor of melon protease is 45700U/g；The additive amount of neutral proteinase is 13g/kg, papain 15g/kg；The temperature of enzymolysis
It spends for 55 DEG C, time 6h.
Compared with prior art, enzymatic hydrolysis and fermentation bone meal of the invention and preparation method thereof, has the advantages that：
Since animal sources selenium-rich product variety is few in the prior art, what the present invention selected is in Ordos of Inner Mongolia
Fermentation bone meal, Se content is made in the ox bone that fanwanxin halal food Co., Ltd of section produces selenium-rich beef with the ox that Se-enriched feedstuff is fed
>=300 micro- gram grams belong to selenium-rich ox bone raw material, add animal sources selenium-rich product variety.
For the first time by neutral proteinase and the double enzyme combination enzymatic hydrolysis and fermentation bovine bone powders of papain, quality improving mitigates bone meal
Granular sensation, bone meal are more smooth, it is determined that enzymolysis and the optimum condition of fermentation impart fermentation after lactobacillus-fermented to bone meal
Flavor has decomposed macromolecular substances, improves flavor and improves nutritional quality, value-added content of product is high, improves the value of ox bone
And utilization rate.
Description of the drawings
Fig. 1 is influence of the hydrolysis temperature to degree of hydrolysis in neutral proteinase experiment of single factor；
Fig. 2 is influence of the time to degree of hydrolysis in neutral proteinase experiment of single factor；
Fig. 3 is that influences of the pH to degree of hydrolysis is digested in neutral proteinase experiment of single factor；
Fig. 4 is that influence of the enzyme additive amount to degree of hydrolysis is digested in neutral proteinase experiment of single factor；
Fig. 5 is influence of the hydrolysis temperature to degree of hydrolysis in papain experiment of single factor；
Fig. 6 is influence of the time to degree of hydrolysis in papain experiment of single factor；
Fig. 7 is that influences of the pH to degree of hydrolysis is digested in papain experiment of single factor；
Fig. 8 is that influence of the enzyme additive amount to degree of hydrolysis is digested in papain experiment of single factor.
Specific embodiment
The specific embodiment of invention is described in detail below, it is to be understood that protection scope of the present invention and from
The limitation of specific embodiment.The test method of actual conditions is not specified in the following example, usually according to normal condition or
According to the condition proposed by each manufacturer.
Embodiment 1
The embodiment of the present invention 1 provides a kind of enzymatic hydrolysis and fermentation bone meal, and the enzymatic hydrolysis and fermentation bone meal is by selenium-rich ox bone raw material
After degreasing fermentation bone meal is made, through neutral proteinase and papain enzymolysis, by lactobacillus bulgaricus and thermophilus
The fermented by mixed bacterium of bacterium, is finally dried to obtain.The preparation method of the enzymatic hydrolysis and fermentation bone meal, comprises the following steps：
S1 chooses selenium-rich ox bone raw material, prepares degreasing fermentation bone meal
The embodiment of the present invention 1 is used using fanwanxin halal food Co., Ltd of section enzymatic hydrolysis and fermentation bone meal in the Inner Mongol, the said firm
Fermentation bone meal is made in the ox bone that the ox that Se-enriched feedstuff is fed produces selenium-rich beef, and fermentation bone meal Se content can reach 300 micrograms,
Belong to enzymatic hydrolysis and fermentation bone meal.Its degreasing fermentation bone meal manufacture craft is as follows：
S11, fermentation bone meal are crude
Selenium-rich ox bone raw material successively through boiling water normal pressure infusion 120min degreasings, 0.1MPa autoclavings 40min with go marrow,
Animal oil, 60-70 DEG C of air-dried 10h, is smashed to pieces, corase grinding, fine grinding, is crossed 100 mesh sieves after grinds, is obtained crude fermentation bone meal；
S12, bone meal degreasing of fermenting
Crude fermentation bone meal is added in the ethyl alcohol that volumetric concentration is 95%, solid-liquid ratio 1g:15ml is stirred in magnetic force
It mixes and is sufficiently stirred degreasing on device (the lofty and steep laboratory apparatus factory in Jintan City west of a city), the time for stirring degreasing is 9h, temperature is 45 DEG C, most
After air-dry, obtain degreasing fermentation bone meal.
S2, enzymolysis
Precise degreasing ferments bone meal in beaker, adds in distilled water, and the bone tankage for being configured to 10g/100ml is molten
Liquid, it is 6 to adjust pH with pH meter, rear to add in neutral proteinase and papain, is placed in constant temperature blender with magnetic force and stirs enzyme
Solution, after digesting, is placed in enzyme deactivation 20min in 90 DEG C of water baths by enzymolysis liquid, obtains enzymatic hydrolysis and fermentation bone meal solution.
Wherein, the vigor of neutral proteinase is 11036U/g, and the vigor of papain is 45700U/g；Neutral proteinase
Additive amount be 13g/kg (using bone tankage solution as mete-wand), papain 15g/kg (using bone tankage solution as
Mete-wand)；The temperature of stirring enzymolysis is 55 DEG C, time 6h.
S3, fermentation
S31 prepares culture medium
Skimmed milk activation culture based formulas：Skimmed milk 100mL, 105 DEG C of sterilizing 5min, it is spare.
Seed liquor is mixed by the component of following mass fraction：Ferment bone meal 15%, glucose 5%, skimmed milk
10%, distilled water 70%；It is packed into bottle, 121 DEG C of 15min that sterilize are spare.
S32, the activation of lactic acid bacteria
Lactobacillus bulgaricus and streptococcus thermophilus are respectively connected in degreasing milk medium, culture 6- in 40.5 DEG C of incubators
10h, until to be placed on 4 DEG C of refrigerators spare for skimmed milk solidification, continuous activation three times, until lactobacillus bulgaricus and streptococcus thermophilus
Cell concentration reach 108-109A/ml respectively obtains the lactobacillus bulgaricus seed liquor of activation and the thermophilus of activation
Bacterium seed liquor.
It should be noted that according to actual amount, using S31 seed liquor respectively the lactobacillus bulgaricus to activation and
The streptococcus thermophilus of activation is enlarged culture, until the lactobacillus bulgaricus seed liquor and thermophilus of volume needed for obtaining
Bacterium seed liquor, and the cell concentration of lactobacillus bulgaricus seed liquor and streptococcus thermophilus seed liquor reaches 108-109A/ml.
S33, inoculation
Add in glucose into enzymatic hydrolysis and fermentation bone meal solution, it is 6 to adjust pH, and sterilizing obtains fermentation medium, then with
The inoculum concentration of 4ml seed liquors/100ml culture mediums, respectively by the lactobacillus bulgaricus seed liquor of activation and the thermophilic chain of activation
Coccus seed liquor is inoculated in fermentation medium, is fermented for 24 hours, obtains fermentation bone meal solution；
Wherein, the usage ratio of enzymatic hydrolysis and fermentation bone meal solution and glucose is 100ml:5g, lactobacillus bulgaricus seed
The volumetric usage ratio of liquid and streptococcus thermophilus seed liquor is 2:1；
Fermentation bone meal solution is evaporated (water content≤3% in the fermentation bone meal after being evaporated), seasoning by S4 according to a conventional method
(for addition sodium citrate as flavoring agent, the dosage of sodium citrate accounts for 0.1% of the fermentation bone meal quality after being evaporated), is made enzymolysis
Ferment bone meal, and the chewable tablets of enzymatic hydrolysis and fermentation bone meal is made after tabletting.
Embodiment 2
The embodiment of the present invention 2 provides a kind of enzymatic hydrolysis and fermentation bone meal, and the enzymatic hydrolysis and fermentation bone meal is by selenium-rich ox bone raw material
After degreasing fermentation bone meal is made, through neutral proteinase and papain enzymolysis, by lactobacillus bulgaricus and thermophilus
The fermented by mixed bacterium of bacterium, is finally dried to obtain.The preparation method of the enzymatic hydrolysis and fermentation bone meal, comprises the following steps：
S1 chooses selenium-rich ox bone raw material, prepares degreasing fermentation bone meal
The embodiment of the present invention 2 is used using fanwanxin halal food Co., Ltd of section enzymatic hydrolysis and fermentation bone meal in the Inner Mongol, the said firm
Fermentation bone meal is made in the ox bone that the ox that Se-enriched feedstuff is fed produces selenium-rich beef, and fermentation bone meal Se content can reach 300 micrograms,
Belong to enzymatic hydrolysis and fermentation bone meal.Its degreasing fermentation bone meal manufacture craft is the same as the operating procedure of embodiment 1.
S2, enzymolysis
Precise degreasing ferments bone meal in beaker, adds in distilled water, and the bone tankage for being configured to 10g/100ml is molten
Liquid, it is 8 to adjust pH with pH meter, rear to add in neutral proteinase and papain, is placed in constant temperature blender with magnetic force and stirs enzyme
Solution, after digesting, is placed in enzyme deactivation 20min in 90 DEG C of water baths by enzymolysis liquid, obtains enzymatic hydrolysis and fermentation bone meal solution.
Wherein, the vigor of neutral proteinase is 11036U/g, and the vigor of papain is 45700U/g neutral proteinases
Additive amount be 13g/kg, papain 15g/kg；The temperature of stirring enzymolysis is 55 DEG C, time 6h.
S3, fermentation
S31 prepares culture medium
Skimmed milk activation culture based formulas：Skimmed milk 100mL, 105 DEG C of sterilizing 5min, it is spare.
Seed liquor is mixed by the component of following mass fraction：Ferment bone meal 15%, glucose 5%, skimmed milk
10%, distilled water 70%；It is packed into bottle, 121 DEG C of 15min that sterilize are spare.
S32, the activation of lactic acid bacteria
Lactobacillus bulgaricus and streptococcus thermophilus are respectively connected in degreasing milk medium, culture 6- in 40.5 DEG C of incubators
10h, until to be placed on 4 DEG C of refrigerators spare for skimmed milk solidification, continuous activation three times, until lactobacillus bulgaricus and streptococcus thermophilus
Cell concentration reach 108-109A/ml respectively obtains the lactobacillus bulgaricus of activation and the streptococcus thermophilus of activation.
S33, inoculation
Add in glucose into enzymatic hydrolysis and fermentation bone meal solution, it is 8 to adjust pH, and sterilizing obtains fermentation medium, then with
The inoculum concentration of 3ml seed liquors/100ml culture mediums, respectively by the lactobacillus bulgaricus seed liquor of activation and the thermophilic chain of activation
Coccus seed liquor is inoculated in fermentation medium, carries out fermentation 30h, obtains fermentation bone meal solution；
Wherein, the usage ratio of enzymatic hydrolysis and fermentation bone meal solution and glucose is 100ml:5g, lactobacillus bulgaricus seed
The volumetric usage ratio of liquid and streptococcus thermophilus seed liquor is 2:1；
Fermentation bone meal solution is evaporated (water content≤3% in the fermentation bone meal after being evaporated), seasoning by S4 according to a conventional method
(for addition sodium citrate as flavoring agent, the dosage of sodium citrate accounts for 0.1% of the fermentation bone meal quality after being evaporated), is made enzymolysis
Ferment bone meal, and the chewable tablets of enzymatic hydrolysis and fermentation bone meal is made after tabletting.
Embodiment 3
The embodiment of the present invention 3 provides a kind of enzymatic hydrolysis and fermentation bone meal, and the enzymatic hydrolysis and fermentation bone meal is by selenium-rich ox bone raw material
After degreasing fermentation bone meal is made, through neutral proteinase and papain enzymolysis, by lactobacillus bulgaricus and thermophilus
The fermented by mixed bacterium of bacterium, is finally dried to obtain.The preparation method of the enzymatic hydrolysis and fermentation bone meal, comprises the following steps：
S1 chooses selenium-rich ox bone raw material, prepares degreasing fermentation bone meal
It operates same as Example 1；
S2, enzymolysis
Operate same as Example 1, it is 10g/kg (with bone tankage solution to differ only in the additive amount of neutral proteinase
For mete-wand), papain 16g/kg (using bone tankage solution as mete-wand)；The temperature of stirring enzymolysis is 50 DEG C,
Time is 6h；
S3, fermentation
It operates same as Example 1.
Embodiment 4
The embodiment of the present invention 4 provides a kind of enzymatic hydrolysis and fermentation bone meal, and the enzymatic hydrolysis and fermentation bone meal is by selenium-rich ox bone raw material
After degreasing fermentation bone meal is made, through neutral proteinase and papain enzymolysis, by lactobacillus bulgaricus and thermophilus
The fermented by mixed bacterium of bacterium, is finally dried to obtain.The preparation method of the enzymatic hydrolysis and fermentation bone meal, comprises the following steps：
S1 chooses selenium-rich ox bone raw material, prepares degreasing fermentation bone meal
It operates same as Example 1；
S2, enzymolysis
Operate same as Example 1, it is 14g/kg (with bone tankage solution to differ only in the additive amount of neutral proteinase
For mete-wand), papain 10g/kg (using bone tankage solution as mete-wand)；The temperature of stirring enzymolysis is 53 DEG C,
Time is 5h；
S3, fermentation
It operates same as Example 1.
Embodiment 5
Bone meal enzymolysis process result of the test of fermenting is as follows：
1st, test index assay method
(1) fat content calculates
Soxhlet extraction method measures fat content
X=(M1-M2) × 100
In formula：Fatty content, g/100g in X-sample；
Filter paper bag weight, g before M1-degreasing；
Filter paper bag weight, g after M2-degreasing；
(2) degree of hydrolysis assay method
Ammonia nitrogen is measured using formol titration, and total nitrogen content is analyzed with kjeldahl apparatus
Degree of hydrolysis (%)=(nitrogen pool in total amino acid amount/degreasing fermentation bone meal in hydrolyzate) × 100%
(3) measure of calcium transformation ratio
EDTA methods measure calcium transformation ratio
Calcium transformation ratio (%)=(calcium content in calcium content/degreasing fermentation bone meal in zymotic fluid centrifuged supernatant) × 100%
(4) viable count measures
Hemacytometry measures viable count.
2nd, experimental result
2.1 neutral proteinase single factor test optimum conditions determine
2.1.1 influence of the hydrolysis temperature to degree of hydrolysis
Influence of the hydrolysis temperature to degree of hydrolysis in neutral proteinase experiment of single factor is as shown in Figure 1, as shown in Figure 1, in enzyme
Solve temperature be less than 50 DEG C when, degree of hydrolysis is raised with the rise of temperature, being positively correlated property, this is because the vigor of enzyme with
It the rise of temperature and raises, the protein of hydrolysis gradually increases in the unit interval, so as to which degree of hydrolysis be caused to raise；It is reached in temperature
During to 50-60 DEG C, the vigor of enzyme has reached optimum state, and the stability of enzyme is also higher, at this point, the degree of hydrolysis highest of protein.
When temperature is more than 60 DEG C, the structure of enzyme gradually changes, and the stability of enzyme starts to reduce, and deactivation phenomenom takes place in enzyme,
The activity of enzyme is gradually reduced with the rise of temperature.Therefore, in order to reduce energy consumption, we determined that the single factor experiment neutral protein
The optimum temperature of enzyme enzymatic hydrolysis and fermentation bone meal solution is 50 DEG C.
2.1.2 influence of the time to degree of hydrolysis
Influence of the time to degree of hydrolysis in neutral proteinase experiment of single factor is as shown in Fig. 2, as seen from Figure 2, in water
The solution time is not reaching to before 4h, neutral proteinase and the sufficiently reaction, and being not reaching to full not yet of degreasing fermentation bone meal
The state of sum.Therefore, before this, enzyme-to-substrate can fully react, larger in the hydrolysis rate of this stage enzyme, on enzymatic hydrolyzation
It rises very fast；But as enzyme digestion reaction carries out, reactant concentration continuously decreases, and enzymolysis product gradually increases, and forms more multiple
Object is closed, so as to hinder the progress of enzyme digestion reaction so that enzyme digestion reaction rate gradually slows down, therefore proceeds to 4- in enzyme digestion reaction
During 6h, enzymolysis rate starts to reduce, but degree of hydrolysis is also being continuously increased, and when enzymolysis time reaches 6h, substrate has almost reacted
Entirely, degree of hydrolysis hardly occurs.Therefore, the optimal of bone meal solution we determined that the single factor experiment neutral protease enzymolysis ferment
Time is 6h.
2.1.3 influences of the pH to degree of hydrolysis is digested
Influences of the pH to degree of hydrolysis is digested in neutral proteinase experiment of single factor as shown in figure 3, as seen from Figure 3, when
When digesting pH value between 6-8, the degree of hydrolysis highest of protein in neutral protease enzymolysis fermentation bone meal, when enzymolysis pH is less than 6
When, the degree of hydrolysis of protein is raised with the rise of pH value in the bone meal that ferments, degree of hydrolysis and enzymolysis being positively correlated property of pH value；Work as enzyme
When solving pH more than 8, the degree of hydrolysis of protein is reduced with the rise of pH value in the bone meal that ferments, and degree of hydrolysis is with digesting pH into negative
It closes；This is because when digesting pH not between 6-8, the conformation of neutral proteinase is changed, by enzyme molecule active site
The dissociation of related group, different dissociated states is presented in this pH value condition neutral proteinase and substrate, so that enzymolysis
Reaction is obstructed, and is affected enzyme-to-substrate and is combined generation complex compound, results in the activity reduction of enzyme.Therefore we determined that the single factor test
It is the optimal enzymolysis pH of neutral protease enzymolysis fermentation bone meal solution when pH is 6-8 in experiment.
2.1.4 influence of the enzyme additive amount to degree of hydrolysis is digested
Influence of the enzyme additive amount to degree of hydrolysis is digested in neutral proteinase experiment of single factor as shown in figure 4, can be seen by Fig. 4
Go out, with enzyme additive amount 10g/kg (in every kilogram of bone tankage solution add 10g neutral proteinases, the difference of following enzyme
Additive amount uses this identical metering method) before, with being continuously increased for enzyme additive amount, albumen in degreasing fermentation bone meal
The degree of hydrolysis of matter is constantly rising, and when the additive amount of enzyme reaches 10g/kg, the degree of hydrolysis of substrate starts to be lower, and works as enzyme
Additive amount reach 12g/kg after, the degree of hydrolysis of enzyme is hardly changing.This is because before 10g/kg, substrate
Hydrolysis degree reaches saturation state not yet, and after the additive amount of enzyme reaches 10g/kg, the hydrolysis degree of substrate is gradual
Tend to saturation state, when the additive amount of enzyme reaches 12g/kg, the hydrolysis degree of substrate is fully saturated.Therefore adding in enzyme
After dosage reaches 12g/kg, degree of hydrolysis variation is little, so, we determined that the single factor experiment neutral protease enzymolysis degreasing is sent out
The optimal enzyme additive amount of ferment bone meal solution is 12g/kg.
2.2 papain single factor test optimum conditions determine
2.2.1 influence of the hydrolysis temperature to degree of hydrolysis
Influence of the hydrolysis temperature to degree of hydrolysis as shown in figure 5, exist as seen from Figure 5 in papain experiment of single factor
When less than 60 DEG C, since enzyme activity is relatively low, hydrolysis rate is slower, and the degree of hydrolysis for causing enzyme is relatively low, and degree of hydrolysis is with enzymolysis
The rise of temperature and raise, the being positively correlated property of temperature of degree of hydrolysis and enzyme；With the continuous rising of temperature, when hydrolysis temperature reaches
At 60 DEG C, the degree of hydrolysis of papain reaches maximum, and when temperature is when being gradually increasing, the structure of papain starts to send out
Raw to change, therewith, degree of hydrolysis starts to reduce.So we determined that temperature of single factor experiment papain enzymolysis fermentation bone meal
It spends for 60 DEG C.
2.2.2 influence of the enzymolysis time to degree of hydrolysis
Influence of the enzymolysis time to degree of hydrolysis in papain experiment of single factor as shown in fig. 6, as seen from Figure 6,
When enzymolysis time is less than 4h, also the reaction was complete for the substrate in enzyme, therefore, degree of hydrolysis increase at any time and increase, when
After enzymolysis time reaches 4h, substrate reactions are complete, and the degree of hydrolysis of enzyme starts the state of tending to be steady.Therefore, papain enzymolysis
The optimal enzymolysis time of fermentation bone meal solution is 4h.Degree of hydrolysis is relatively low when reaction starts, with the extension of hydrolysis time, degree of hydrolysis
Increase, but hydrolysis time, after 4h, the change rate of degree of hydrolysis becomes smaller, degree of hydrolysis tends to be constant afterwards.Therefore, papain
Enzymolysis time for 4h it is preferable.
2.2.3 influence of the pH value to degree of hydrolysis is digested
Influence of the pH value to degree of hydrolysis is digested in papain experiment of single factor as shown in fig. 7, as seen from Figure 7,
When pH is 6, degree of hydrolysis highest；When pH is less than 6, the degree of hydrolysis of papain is raised with the rise of pH, when enzymolysis pH is arrived
During up to 6, degree of hydrolysis reaches peak value, and when degree of hydrolysis is less than 6, the degree of hydrolysis of papain is reduced with the rise of pH value.When
During higher or lower than 6, papain is combined with the protein in fermentation bone meal solution and can be obstructed, so as to cause pawpaw egg
The decline of white enzymatic activity or even deactivation, so choosing the optimal enzyme that pH 6 is papain enzymolysis fermentation bone meal solution
Solve pH.
2.2.4 influence of the enzyme additive amount to degree of hydrolysis is digested
Influence of the enzyme additive amount to degree of hydrolysis is digested in papain experiment of single factor as shown in figure 8, can be seen by Fig. 8
Go out, before enzyme additive amount is 10g/kg, degree of hydrolysis rapid increase with the increase of enzyme additive amount, when the additive amount of enzyme reaches
During 10g/kg, the increase of degree of hydrolysis starts slack-off, and when enzyme additive amount is 14g/kg, the degree of hydrolysis of enzyme reaches maximum.
So select optimum additions of the enzyme additive amount 15g/kg for papain enzymolysis fermentation bone meal solution.
2.3 enzymatic hydrolysis conditions screen orthogonal test
By above-mentioned 2.1 and 2.2 single factor experiment, the optimum condition for obtaining neutral proteinase single factor test is：50 DEG C, enzyme
Time 6h, pH 6-8 are solved, enzyme additive amount is 12g/kg.The optimum condition of papain single factor test is：60 DEG C, enzymolysis time
For 4h, pH 6, enzymolysis dosage 15g/kg.When designing orthogonal test, enzymolysis time is set as that 6h and enzymolysis pH are set to definite value 6,
A neutral protease enzymolysis temperature (DEG C), B be neutral proteinase enzyme dosage (g/kg), C be papain enzymolysis temperature (DEG C), D
For papain enzyme additive amount (g/kg), it is as shown in table 1 that fermentation bone meal digests Orthogonal Experiment and Design scheme.
The fermentation bone meal enzymolysis Orthogonal Experiment and Design scheme of table 1
The data such as table 2 measured after said program enzymolysis.As it can be seen from table 1 each factor influence order is followed successively by B>D
>A>C, i.e. neutral protease enzymolysis temperature>Papain enzymolysis temperature>Neutral proteinase additive amount>Papain adds
Amount.Optimum technological parameter is the A2B2C3D1 in table 1, that is, chooses the enzyme dosage of neutral proteinase as 13g/kg, neutral egg
The hydrolysis temperature of white enzyme is 55 DEG C, and the enzyme dosage of papain is 15g/kg, and the hydrolysis temperature of papain is 55 DEG C.
Orthogonal test shows the degree of hydrolysis for being greater than single enzyme hydrolysis with the degree of hydrolysis of two kinds of enzyme hydrolysis, such as：In pawpaw
Protease and neutral proteinase, hydrolysis temperature are 60 DEG C, and enzymolysis time is 6h, and enzymolysis pH value is 6, and enzyme additive amount is
In the case of 13g/kg；The degree of hydrolysis of neutral proteinase is 6.12%, and the degree of hydrolysis of papain is 6.78%, and two kinds enzyme-linked
The degree of hydrolysis of Heshui solution is 10.21%.Therefore, double-enzyme hydrolysis is better than single enzymolysis to a certain extent.
The orthogonal result table of 2 proteolysis assay of table
Therefore we determined that, fermentation bone meal optimum hydrolysising condition is：Bone meal concentration of fermenting 10g/100ml；Double-enzyme hydrolysis；Wood
The hydrolysis temperature of melon protease is 55 DEG C, and enzyme additive amount is 15g/kg, and enzymolysis pH value is 6, enzymolysis time 4h；Neutral proteinase
Hydrolysis temperature for 55 DEG C, enzyme additive amount is 13g/kg, and enzymolysis pH value is 6, enzymolysis time 6h；The optimal fermentation of fermentation bone meal
Condition is：Bone meal concentration of fermenting 10g/100ml；The strain ratio of lactobacillus bulgaricus and streptococcus thermophilus is 2:1；Most preferably connect
Kind amount is 3%；Glucose additive amount is 3%, fermentation time 30h；The water of double-enzyme hydrolysis under the hydrolysising condition and fermentation condition
Solution rate is 12.58%；The conversion ratio of calcium is 31.45%.
It should be noted that involved in claims of the present invention during numberical range, it is thus understood that each numberical range
Any one numerical value can be selected between two endpoints and two endpoints, due to step method and the embodiment 1-4 phases of use
Together, repeat in order to prevent, the present invention describes preferred embodiment 1-2 and its effect, but those skilled in the art once obtain
Cicada basic creative concept can then make these embodiments other change and modification.So appended claims are intended to
It is construed to include preferred embodiment and falls into all change and modification of the scope of the invention.
Obviously, various changes and modifications can be made to the invention without departing from essence of the invention by those skilled in the art
God and scope.In this way, if these modifications and changes of the present invention belongs to the scope of the claims in the present invention and its equivalent technologies
Within, then the present invention is also intended to comprising including these modification and variations.
Claims (6)
1. a kind of enzymatic hydrolysis and fermentation bone meal, which is characterized in that the enzymatic hydrolysis and fermentation bone meal is that selenium-rich ox bone raw material is made degreasing hair
After ferment bone meal, through neutral proteinase and papain enzymolysis, by lactobacillus bulgaricus and the Mixed Microbes of streptococcus thermophilus
Kind fermentation, is finally dried to obtain.
2. a kind of preparation method of enzymatic hydrolysis and fermentation bone meal described in claim 1, which is characterized in that comprise the following steps：
S1 chooses selenium-rich ox bone raw material, prepares degreasing fermentation bone meal；
S2, enzymolysis
The bone tankage solution of 10g/100ml is prepared, adjusting pH value is 6-8, rear to add in neutral proteinase and papain, enzyme
Solution, after digesting, is placed in 90 DEG C of enzyme deactivation 20min by enzymolysis liquid, obtains enzymatic hydrolysis and fermentation bone meal solution；
Wherein, the additive amount of neutral proteinase is 10-14g/kg, and papain 10-16g/kg, hydrolysis temperature is 50-55 DEG C,
Enzymolysis time is 4-6h；
S3, fermentation
Glucose is added in into enzymatic hydrolysis and fermentation bone meal solution, adjusting pH value is 6-8, is sterilized, the guarantor for being then respectively adding activation adds
Leah lactobacillus seed liquor and the streptococcus thermophilus seed liquor of activation, carry out fermentation 24-30h, obtain fermentation bone meal solution；
Wherein, the usage ratio of enzymatic hydrolysis and fermentation bone meal solution and glucose is 100ml:5g, lactobacillus bulgaricus seed liquor and
The volumetric usage ratio of streptococcus thermophilus seed liquor is 2:1；
Fermentation bone meal solution is evaporated, seasons, enzymatic hydrolysis and fermentation bone meal is made by S4.
3. the preparation method of enzymatic hydrolysis and fermentation bone meal according to claim 2, which is characterized in that prepare degreasing fermentation bone meal
Technique is as follows：
S11, fermentation bone meal are crude
Selenium-rich ox bone raw material through degreasing, removes marrow, animal oil successively, air-dries, smashs to pieces, sieve after grinds, obtain crude fermentation
Bone meal；
S12, bone meal degreasing of fermenting
Crude fermentation bone meal is added in the ethyl alcohol that volumetric concentration is 95%, solid-liquid ratio 1g:15ml, degreasing, last wind
It is dry, obtain degreasing fermentation bone meal.
4. the preparation method of enzymatic hydrolysis and fermentation bone meal according to claim 3, which is characterized in that S11's concretely comprises the following steps：It is rich
For selenium ox bone raw material successively through boiling water normal pressure infusion 120min degreasings, 0.1MPa autoclaving 40min remove marrow, animal oil, 60-70
DEG C air-dried 10h, is smashed to pieces, and 100 mesh sieves are crossed after grinds, obtain crude fermentation bone meal.
5. the preparation method of enzymatic hydrolysis and fermentation bone meal according to claim 3, which is characterized in that in S12, the time of degreasing is
9h, temperature are 45 DEG C.
6. the preparation method of enzymatic hydrolysis and fermentation bone meal according to claim 3, which is characterized in that in S2, neutral proteinase
Vigor is 11036U/g, and the vigor of papain is 45700U/g；The additive amount of neutral proteinase be 13g/kg, Papain
Enzyme 15g/kg；The temperature of enzymolysis is 55 DEG C, time 6h.
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| CN109549215A (en) * | 2018-11-23 | 2019-04-02 | 内蒙古神元康肽生物工程有限公司 | The preparation method of one breeding ass bone collagen peptide oral solution |
| CN110663883A (en) * | 2018-10-26 | 2020-01-10 | 福建省亚明食品有限公司 | Method for improving yield of spare rib dish by multiple times of microbial fermentation |
| CN113801909A (en) * | 2021-10-14 | 2021-12-17 | 内蒙古农业大学 | Preparation method and application of sheep bone calcium binding peptide |
| CN114982895A (en) * | 2022-05-25 | 2022-09-02 | 石河子大学 | Fermented bovine bone meal prebiotics effervescent tablet and preparation method thereof |
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| CN102318794A (en) * | 2011-10-19 | 2012-01-18 | 郭景龙 | Health-care food having calcium supplementing function |
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| CN102318794A (en) * | 2011-10-19 | 2012-01-18 | 郭景龙 | Health-care food having calcium supplementing function |
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN110663883A (en) * | 2018-10-26 | 2020-01-10 | 福建省亚明食品有限公司 | Method for improving yield of spare rib dish by multiple times of microbial fermentation |
| CN109549215A (en) * | 2018-11-23 | 2019-04-02 | 内蒙古神元康肽生物工程有限公司 | The preparation method of one breeding ass bone collagen peptide oral solution |
| CN113801909A (en) * | 2021-10-14 | 2021-12-17 | 内蒙古农业大学 | Preparation method and application of sheep bone calcium binding peptide |
| CN114982895A (en) * | 2022-05-25 | 2022-09-02 | 石河子大学 | Fermented bovine bone meal prebiotics effervescent tablet and preparation method thereof |
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