CN102326796B - Method for producing oyster hydrolyzate through microbial fermentation - Google Patents
Method for producing oyster hydrolyzate through microbial fermentation Download PDFInfo
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- CN102326796B CN102326796B CN2011101717925A CN201110171792A CN102326796B CN 102326796 B CN102326796 B CN 102326796B CN 2011101717925 A CN2011101717925 A CN 2011101717925A CN 201110171792 A CN201110171792 A CN 201110171792A CN 102326796 B CN102326796 B CN 102326796B
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Abstract
The invention discloses a method for preparing an oyster hydrolyzate through microbial fermentation, which is characterized in that a pacific oyster is fermented under the action of Aspergillus oryzae to prepare an oyster hydrolyzate. The preparation process comprises the following steps: taking fresh oyster meat, cleaning, homogenizing, and inoculating with the Aspergillus oryzae for fermentation; and after the fermentation is finished, carrying out filtration sterilization, and concentrating the filtrate to obtain the oyster hydrolyzate. By adopting a liquid fermentation mode, the method has the advantages of reasonable design of process parameters, high operability, low cost and high total nitrogen extraction rate and glycogen extraction rate, avoids the use of finished proteases and amylases, can make full use of oyster resources, and has better industrial application potential.
Description
Technical field
The present invention relates to the method that a kind of liquid fermentation oyster prepares oyster hydrolyzate, belong to processing of aquatic products and field of microbial biotechnology.
Background technology
The oyster delicious flavour, nutritious, have the good reputation of " marine milk ", be that China classifies one of health care curative effect product of integration of drinking and medicinal herbs in the first batch as.Oyster Protein is higher up to the content of 50%, 8 kind of essential amino acid, accounts for 40% of total amino acid content, and required histidine and the arginine content of infant is higher, is a kind of good protein.Contain the taurine more than 3% in the what is more important oyster meat.Taurine has the effects such as eliminating inflammation and expelling toxin, hepatic cholagogic, reducing blood lipid, promotion child brain development and Anshen Bunao.In addition, also contain in the oyster meat up to 22.41% have mineral matter and trace elements such as unrighted acid, vitamin and iron, zinc, calcium, manganese, selenium such as important physiologically active glycogen, DHA (DHA) and eicosapentaenoic acid (EPA).
Oyster is one of China's four large cultivated shellfishes, propagates in a large number aboundresources artificially in the coastal area.At present, oyster processing and comprehensive utilization are also relatively backward, are main mainly with eating and make the biltong goods raw, are processed on a small quantity oyster sauce or other flavouring.The oyster deep processing prepares high value added product and rarely has report, and the phenomenon of " Higher output is not accompanied by a higher income " appears in therefore domestic oyster culture industry.Generation is easy to absorb behind the proteolysis, dissolubility is good, emulsibility is good, and has the polypeptide of multiple physiologically active, has improved the functional characteristic of protein, has improved nutrition and the value of protein.
Pacific oyster claims again long oyster, Japan real oyster, wide warm euryhalinous inner bay kind.At present, the China coast each province cultivates this kind more.Enzymolysis property and the technique that comprises the multiple oysters such as Pacific oyster, Crassostrea rivularis, ostrea talienwhanensis Crosse is studied both at home and abroad at present.With oyster hydrolysis, can produce and have the active peptides that comprises antifatigue, the multiple physiologically active such as anti-oxidant.Report comprises that the oyster hydrolysis all is directly to add the finished product protease hydrolytic at present.The finished product protease price is expensive, has increased production cost.
Summary of the invention
Technical problem to be solved by this invention is for the deficiencies in the prior art, provide a kind of new technique more rationally, effectively reduce cost, applicable to the method for the producing oyster hydrolyzate through microbial fermentation of the large production of industry.
Technical problem to be solved by this invention is to realize by following technical scheme.The present invention is a kind of method of producing oyster hydrolyzate through microbial fermentation, is characterized in, it adopts aspergillus oryzae Aspergillus oryzae liquid fermentation method, and its concrete steps are as follows:
(1) preparation of oyster: buy fresh Pacific oyster, clean up, take out oyster meat, refiner homogenate;
(2) preparation fermentation substrate: homogenate is 15% with distilled water diluting to the M/V of protein concentration, and with the NaOH adjusting pH value to 7.0 of 1M, 60 ℃ of pasteurization 30min get fermentation substrate;
(3) microbial fermentation: aspergillus oryzae Aspergillus oryzae bacterial classification is activated at the PDA slant medium, and the picking spore inoculating is to the PDA fluid nutrient medium on the culture presevation inclined-plane after the activation, and 30 ℃, 120rpm are cultured to cell concentration and reach 10
7CFU/ml gets the fermentation strain seed liquor; By V/V 5% inoculum concentration the fermentation strain seed liquor is inoculated in fermentation substrate, liquid amount V/V 20%, rotating speed 140rpm, 30 ℃, fermentation 48h;
(4) separate: after the fermentation ends, remove thalline, not protein hydrolysate and other impurity with the milipore filter ultrafiltration of holding back 100KD, cleaner liquid is the oyster hydrolysate, perhaps the powder process of cleaner liquid concentrate drying is got powdery oyster hydrolysate.
Potato culture (PDA) can adopt the following methods configuration: potato 200g (section poach 30min crosses leaching juice), sucrose 20.0g, distilled water 1000mL, pH value nature.It is 2% agar that solid medium adds m/V in addition.
The aspergillus oryzae Aspergillus oryzae (bacterial strain AS3.951) that can select available from Institute of Micro-biology of the Chinese Academy of Sciences of the present invention.
Below be the inventor do about the technical scheme of the inventive method or optimization test and the result thereof of parameter.
1, the preparation of working sample: respectively in the sampling of different fermentation time, boil the 10min enzyme that goes out.Zymotic fluid is crossed 0.45 μ m miillpore filter, and filtrate is measured as sample.
2, inoculum concentration is on the impact of total nitrogen recovery rate and glycogen recovery rate:
After preparing fermentation substrate according to the inventive method step (2), inoculum concentration is respectively 2.5%, 5%, 10%, 15%, liquid amount is V/V 20%, rotating speed 120rpm, 30 ℃ of fermented and cultured are respectively 0,12,24,36,48,60,72,84,96h sampling is measured inoculum concentration to the impact of total nitrogen recovery rate and glycogen recovery rate.Result such as Fig. 1, inoculum concentration variable effect total nitrogen recovery rate and glycogen recovery rate, when inoculum concentration was 2.5%, the total nitrogen recovery rate reached maximum at fermentation 60h, and inoculum concentration all reaches the higher extracted level at fermentation 48h when being 5%, 10%, 15%; When inoculum concentration was 2.5% and 5%, the glycogen recovery rate reached the higher extracted level at fermentation 72h and 48h respectively, and inoculum concentration is when being 10% and 15%, and the two all reaches the higher extracted level about fermentation 36h.Comprehensive total nitrogen recovery rate, glycogen recovery rate and cost, inoculum concentration selects 5%, and this condition bottom fermentation 48h total nitrogen recovery rate and glycogen recovery rate all reach the higher extracted level.
3, initial pH is on the impact of total nitrogen recovery rate and glycogen recovery rate:
With distilled water homogenate is diluted to protein concentration m/V 15%, is respectively 5.0,6.0,7.0,8.0,9.0,60 ℃, the 30min pasteurization with 1M NaOH or 1M salt acid for adjusting pH value.Inoculum concentration V/V 5%, liquid amount V/V 20%, and rotating speed 120rpm, 30 ℃ of fermented and cultured in 0,12,24,36,48,60,72,84,96h sampling, are measured initial pH to the impact of total nitrogen recovery rate and glycogen recovery rate respectively.Result such as Fig. 2, the total nitrogen recovery rate reaches maximum the earliest when pH8, and glycogen recovery rate recovery rate when pH6 is the highest, and reach the earliest maximum, and balance is considered protein extracting ratio and glycogen recovery rate, selecting the fermentation initial pH value is 7.0.
4, concentration of substrate is on the impact of total nitrogen recovery rate and glycogen recovery rate:
With distilled water homogenate is diluted to protein concentration m/V 5%, 10%, 15%, 20%, regulates pH value to 7.0,60 ℃, 30min pasteurization with 1M NaOH.Inoculum concentration V/V5%, liquid amount V/V 20%, rotating speed 120rpm, 30 ℃ of fermented and cultured in 0,12,24,36,48,60,72,84,96h sampling, are measured initial protein concentration to the impact of total nitrogen recovery rate and glycogen recovery rate respectively.Result such as Fig. 3, not remarkable on the impact of total nitrogen recovery rate and glycogen recovery rate when protein concentration is 5%, 10% and 15%, but therefore total nitrogen recovery rate and all significantly decline of glycogen recovery rate when protein concentration is 20% select protein concentration 15%.
5, fermentation temperature is on the impact of total nitrogen recovery rate and glycogen recovery rate:
With distilled water homogenate is diluted to protein concentration m/V 15%, regulates pH value to 7.0,60 ℃, 30min pasteurization with 1M NaOH.Inoculum concentration V/V 5%, liquid amount V/V 20%, and rotating speed 120rpm, respectively at 26 ℃, 28 ℃, 30 ℃, 32 ℃ of fermented and cultured respectively in 0,12,24,36,48,60,72,84,96h sampling, are measured fermentation temperature to the impact of total nitrogen recovery rate and glycogen recovery rate.Result such as Fig. 4, temperature total nitrogen recovery rate and glycogen recovery rate in the time of 30 ℃ reach peak the earliest, and therefore selecting fermentation temperature is 30 ℃.
6, liquid amount is on the impact of total nitrogen recovery rate and glycogen recovery rate:
With distilled water homogenate is diluted to protein concentration m/V 15%, regulates pH value to 7.0,60 ℃, 30min pasteurization with 1M NaOH.Inoculum concentration V/V 5%, and liquid amount is adjusted into respectively V/V 10%, 20%, 30%, 40%, rotating speed 120rpm, and 30 ℃ of fermented and cultured are respectively 0,12,24,36,48,60,72,84,96h sampling is measured liquid amount to the impact of total nitrogen recovery rate and glycogen recovery rate.Result such as Fig. 5, total nitrogen recovery rate and glycogen recovery rate reached peak the earliest when liquid amount was 20%, and therefore selecting liquid amount is 20%.
7, rotating speed is on the impact of total nitrogen recovery rate and glycogen recovery rate:
With distilled water homogenate is diluted to protein concentration m/V 15%, regulates pH value to 7.0,60 ℃, 30min pasteurization with 1M NaOH.Inoculum concentration V/V 5%, and liquid amount V/V 20%, rotating speed are set as respectively 100rpm, 120rpm, 140rpm, 160rpm, and 30 ℃ of fermented and cultured are respectively 0,12,24,36,48,60,72,84,96h sampling is measured rotating speed to the impact of total nitrogen recovery rate and glycogen recovery rate.Result such as Fig. 6, total nitrogen recovery rate and glycogen recovery rate reached peak the earliest when rotating speed was 140rpm, and therefore selecting rotating speed is 140rpm.
8, fermentation time is on the impact of total nitrogen recovery rate, glycogen recovery rate, protease, amylase activity:
With distilled water homogenate is diluted to protein concentration m/V 15%, regulates pH value to 7.0,60 ℃, 30min pasteurization with 1M NaOH.Inoculum concentration V/V 5%, liquid amount V/V 20%, and rotating speed 140rpm, 30 ℃ of fermented and cultured respectively in 0,12,24,36,48,60,72,84,96h sampling, are measured fermentation time to the impact of total nitrogen recovery rate, glycogen recovery rate, proteinase activity, amylase activity.Result such as Fig. 7, along with the prolongation of fermentation time, total nitrogen recovery rate, glycogen recovery rate, proteinase activity, amylase activity increase, and total nitrogen recovery rate, glycogen extraction rate reached are to maximum behind the fermentation 48h, and therefore selecting fermentation time is 48h.
9, hydrolyzate separates and measures: after the fermentation ends, can remove thalline and not protein hydrolysate and other impurity with the U.S. Millipore company milipore filter ultrafiltration of holding back 100KD.Measure free amino acid and the taurine of ultrafiltrate and find that the ultrafiltrate free amino acid accounts for 35.91% of total amino acid, taurine accounts for 13.05% of total amino acid.
Compared with prior art, the inventive method adopts the liquid fermentation form, and its process parameters design is reasonable, workable, need not to use finished product protease and amylase, cost is low, total nitrogen recovery rate and glycogen recovery rate are high, can take full advantage of the oyster resource, have preferably commercial Application potentiality.
Description of drawings
Fig. 1 is that inoculum concentration is on the figure that affects of total nitrogen recovery rate and glycogen recovery rate; Among the figure: ■, ●, ▲,
Being respectively inoculum concentration is 2.5%, 5%, total nitrogen recovery rate in the time of 10%, 15%; , zero, △,
Being respectively inoculum concentration is 2.5%, 5%, glycogen recovery rate in the time of 10%, 15%;
Fig. 2 is that initial pH is on the figure that affects of total nitrogen recovery rate and glycogen recovery rate; Among the figure: ■, ●, ▲,
◆ being respectively initial pH is 5,6,7,8,9 o'clock total nitrogen recovery rates; , zero, △,
, it is 5,6,7,8 that ◇ is respectively initial pH, 9 o'clock glycogen recovery rates;
Fig. 3 is that concentration of substrate is on the figure that affects of total nitrogen recovery rate and glycogen recovery rate; Among the figure: ■, ●, ▲,
Being respectively concentration of substrate is 5%, 10%, total nitrogen recovery rate in the time of 15%, 20%; , zero, △,
Being respectively concentration of substrate is 5%, 10%, glycogen recovery rate in the time of 15%, 20%; Fig. 4 is that fermentation temperature is on the figure that affects of total nitrogen recovery rate and glycogen recovery rate; Among the figure: ■, ●, ▲,
Being respectively fermentation temperature is 26 ℃, 28 ℃, and 30 ℃, total nitrogen recovery rate in the time of 32 ℃; , zero, △,
Being respectively fermentation temperature is 26 ℃, 28 ℃, and 30 ℃, glycogen recovery rate in the time of 32 ℃
Fig. 5 is that liquid amount is on the figure that affects of total nitrogen recovery rate and glycogen recovery rate; Among the figure: ■, ●, ▲,
Being respectively liquid amount is 10%, 20%, total nitrogen recovery rate in the time of 30%, 40%; , zero, △,
Being respectively liquid amount is 10%, 20%, glycogen recovery rate in the time of 30%, 40%;
Fig. 6 is that rotating speed is on the figure that affects of total nitrogen recovery rate and glycogen recovery rate; Among the figure: ■, ●, ▲,
Being respectively rotating speed is 100rpm, 120rpm, 140rpm, total nitrogen recovery rate during 160rpm; , zero, △,
Being respectively rotating speed is 100rpm, 120rpm, 140rpm, glycogen recovery rate during 160rpm;
Fig. 7 is that fermentation time produces protease and the diastatic figure that affects to protein recovery, glycogen recovery rate and aspergillus oryzae; Among the figure: the ■ protein recovery; ● the glycogen recovery rate; ▲ proteinase activity;
Amylase activity.
The specific embodiment
Below further describe concrete technical scheme of the present invention, so that those skilled in the art understands the present invention further, and do not consist of its Copyright law.
Embodiment 1, and a kind of microbial fermentation prepares the method for oyster hydrolysate, and it adopts aspergillus oryzae Aspergillus oryzae liquid fermentation method, and its concrete steps are as follows:
(1) preparation of oyster: buy fresh Pacific oyster, clean up, take out oyster meat, refiner homogenate;
(2) preparation fermentation substrate: homogenate is 15% with distilled water diluting to the M/V of protein concentration, and with the NaOH adjusting pH value to 7.0 of 1M, 60 ℃ of pasteurization 30min get fermentation substrate;
(3) microbial fermentation: aspergillus oryzae Aspergillus oryzae bacterial classification is activated at the PDA slant medium, from the activation after the culture presevation inclined-plane on the picking spore inoculating to the PDA fluid nutrient medium, 30 ℃, 120rpm are cultured to cell concentration and reach 107CFU/ml, get the fermentation strain seed liquor; By V/V 5% inoculum concentration the fermentation strain seed liquor is inoculated in fermentation substrate, liquid amount V/V 20%, rotating speed 140rpm, 30 ℃, fermentation 48h; (4) separate: after the fermentation ends, remove thalline, not protein hydrolysate and other impurity with the milipore filter ultrafiltration of holding back 100KD, cleaner liquid is the oyster hydrolysate, perhaps the powder process of cleaner liquid concentrate drying is got powdery oyster hydrolysate.
Claims (1)
1. a microbial fermentation prepares the method for oyster hydrolysate, and it is characterized in that: it adopts aspergillus oryzae
Aspergillus oryzaeLiquid fermentation method, its concrete steps are as follows:
(1) preparation of oyster: buy fresh Pacific oyster, clean up, take out oyster meat, refiner homogenate;
(2) preparation fermentation substrate: homogenate is 15% with distilled water diluting to the M/V of protein concentration, and with the NaOH adjusting pH value to 7.0 of 1M, 60 ℃ of pasteurization 30min get fermentation substrate;
(3) microbial fermentation: with aspergillus oryzae
Aspergillus oryzaeBacterial classification activates at the PDA slant medium, and the picking spore inoculating is to the PDA fluid nutrient medium on the culture presevation inclined-plane after the activation, and 30 ℃, 120rpm are cultured to cell concentration and reach 10
7CFU/ml gets the fermentation strain seed liquor; By V/V 5% inoculum concentration the fermentation strain seed liquor is inoculated in fermentation substrate, liquid amount V/V 20%, rotating speed 140rpm, 30 ℃, fermentation 48h;
(4) separate: after the fermentation ends, remove thalline, not protein hydrolysate and other impurity with the milipore filter ultrafiltration of holding back 100KD, cleaner liquid is the oyster hydrolysate, perhaps the powder process of cleaner liquid concentrate drying is got powdery oyster hydrolysate.
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| CN105861565B (en) * | 2016-06-04 | 2019-08-23 | 中国海洋大学 | A kind of turbot fish-skin tunning with antioxidant activity |
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| CN102787079B (en) * | 2012-08-06 | 2014-09-17 | 江南大学 | Aspergillus oryzae and application method thereof in preparing sauce fragrant flavor material |
| CN104172080B (en) * | 2014-08-05 | 2015-11-18 | 北海富安源生物科技有限公司 | Amino acid whose method is extracted in oyster meat |
| CN108396048A (en) * | 2018-02-08 | 2018-08-14 | 舟山海研食品科技有限公司 | The preparation method of oyster active peptides |
| KR102132862B1 (en) * | 2018-06-15 | 2020-07-10 | 제주대학교 산학협력단 | Composition for bone health comprising functional fermented material using oyster |
| CN109401983B (en) * | 2018-11-20 | 2020-01-10 | 佛山市海天(高明)调味食品有限公司 | Aspergillus oryzae ZA151 and application thereof |
| CN115777902B (en) * | 2022-12-07 | 2024-08-06 | 佛山市海天(宿迁)调味食品有限公司 | Composite flavor oyster juice, oyster oil and preparation method thereof |
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| CN105861565B (en) * | 2016-06-04 | 2019-08-23 | 中国海洋大学 | A kind of turbot fish-skin tunning with antioxidant activity |
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