CN103300210B - Method for preparing compound enzyme and probiotic preparation for feed by biotransformation of reed and peanut cake - Google Patents
Method for preparing compound enzyme and probiotic preparation for feed by biotransformation of reed and peanut cake Download PDFInfo
- Publication number
- CN103300210B CN103300210B CN201310229627.XA CN201310229627A CN103300210B CN 103300210 B CN103300210 B CN 103300210B CN 201310229627 A CN201310229627 A CN 201310229627A CN 103300210 B CN103300210 B CN 103300210B
- Authority
- CN
- China
- Prior art keywords
- reed
- feed
- fermentation
- peanut dregs
- enzyme
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Expired - Fee Related
Links
Abstract
The invention discloses a method for preparing a compound enzyme and probiotic preparation for feed by biotransformation of reed and peanut cake. The method comprises the following steps of: fully mixing Pu'er ripe tea, shiitake mushroom log, agrocybe aegerita mushroom log, first reed, first peanut cake, first wheat bran, first apple pomace and first water which are fermented by piling, and performing acclimation culture so as to form a fermentation agent; and fully mixing the fermentation agent, second reed, second peanut cake, second wheat bran, second apple pomace and second water, and fermenting so as to obtain the compound enzyme and probiotic preparation for feed. In the method disclosed by the invention, a food-grade safe bacterial group with stable microecology is utilized for fermenting the reed and the peanut cake so as to prepare the compound enzyme and probiotic preparation for feed, which has various better performance indexes, and the low-cost and high-efficient biotransformation of the reed and the peanut cake can be realized. Simultaneously, the method disclosed by the invention has the advantages of easiness in growth of microbes, high enzyme activity, extensive fermentation process and easiness in industrial large-scale production.
Description
Technical field
The invention belongs to field of microbial fermentation, be specifically related to a kind of grade-safe flora fermentation reed and peanut dregs adopting Tiny ecosystem stable, prepare complex enzyme for feed and to hold concurrently the method for probiotics preparation.
Background technology
China is the populous nation that an arable land puts upon the full stretch, and the demand of feedstuff particularly energy feed and protein feed all exists huge breach.
Lignocellulosic is renewable resource abundant, the most cheap on the earth.The process of manufacture of agricultural product and food also produces a large amount of lignocellulose accessory substance.Lignocellulosic is of a great variety, such as various agricultural crop straw, city cellulosic rubbish, processing of farm products accessory substance are as corncob, vegetable waste, fruits and vegetables slag etc., the composition of lignocellulosic mainly comprises cellulose, hemicellulose, lignin and pectin etc., the complex substrate that occurring in nature has major application value is become because of its large cheapness, but due to composition and the complex structure of lignocellulosic, be thus difficult to by simple stomach animal digestion.Realize the non-grain material high efficiency, low cost biologies such as lignocellulosic to transform, making the cereal such as its replacement of corn as energy feed, will be the important growth point of Feed Industry Development.
Grouts are the rich proteinaceous large agricultural by-products of a class.Along with the fast development of aquaculture industry, also increasing to protein resource demand, and animality high-quality protein resource (as fish meal) limited source, price is in recent years always in quick rise, therefore, take grouts as protein feed, greatly will alleviate the nervous situation of feed protein resource, and can feed cost be reduced.Grouts are of a great variety, comprise soybean cake dregs, cottonseed (benevolence) grouts, rapeseed cake dregs, peanut (benevolence) grouts, sunflower (benevolence) grouts, teaseed cake dregs, sesame cake meal etc., but containing very various toxicant and anti-nutrient substance in grouts.Such as sulphur glycosides decomposites the noxious material such as isothiocyanates and nitrile through myrosase having under regimen condition, makes animal Thyroid Gland Swell, metabolic disturbance, even causes subcutaneous hemorrhage, also affect the function of the organs such as adrenal cortex, pituitary and liver.Anti-nutrient substance alpha-galactoside compound then makes animal intestinal flatulence thus affects digestive function.And grouts also contain quite high plant fiber component richness is proteinaceous while, nonruminant digestion not easily.If toxicity in grouts and anti-nutrient substance can be eliminated and carry out biology conversion in high efficiency, low cost ground, then all kinds of grouts will be developed as protein feed raw material and the high price such as Peru Fish Dietary animal fodder albumen fully, become another important growth point of future in feed industry development.
Fermentable is the effective way realizing lignocellulosic and puffed soybean agricultural by-products low-cost high-efficiency biotransformation.Lignocellulosic is carbohydrate substantially, grouts are rich in protein, both combinations can be microorganism provides nutritious, well-proportioned fermentation substrate, and system biological theory is then for the low-cost high-efficiency biotransformation being realized lignocellulosic and grouts by fermentable provides New Policy.According to system biological theory, microbiologic population and its enzyme system and substrate environment residing for it are indivisible holonomic systems, and it is formed is the result of long-term natural evolution and balance.In such a system, different microorganisms produce the kind of enzyme and vigor is had nothing in common with each other, only to coexist in same system and the environmental factor that provides of shared system, could effective conversion of substrate.In the wild microbiologic population of the Tiny ecosystem stable equilibrium that occurring in nature ubiquity is made up of many kind different microorganisms, such as rumen microorganism group, biogas microbiologic population, compost microbe group, feed ensiling microbiologic population, higher mammal intestinal microflora, endophyte of plant group, white-rot fungi group, xylophaga enteron aisle endophyte group etc.These wild micropopulations are dropped in specific wild environment residing for it can carry out stable mixed culture fermentation, and due to its produce the diversity of enzyme system, biology can be carried out to complex substrate and transform.Therefore, according to system biological theory, by on the basis of natural flora, build one and produce enzyme and powerful and applicable lignocellulosic and grouts matrix environment the Tiny ecosystem of fermentability and stablize flora, be expected the low-cost high-efficiency biotransformation realizing lignocellulosic and puffed soybean.
Pu'er tea is a kind of fermented tea, and it defines the complicated mutually but flora that Tiny ecosystem is stable of bacterium in pile-fermentation process, i.e. the wet heap flora of Pu'er tea, and this flora comprises aspergillus (Aspergiltus), rhizopus (Rhizopus), Penicillium (Penicillium), trichoderma (Trichoderma), Candida parapsilosis (C.parapsilosis), candida famata (C.famata), candida ciferrii (C.cifferrii), Crewe takes yeast (Saccharomyces kluyoerii), Cryptococcus laurentii (Cryptococcus laurentii), basidiomycetes (Basidiomycetes), lactobacillus (Lactobacillus), bacillus (Bacillus), brevibacterium (Brevibaterium), Coccus (Staphylococcus), line Pseudomonas (Actinoplanes), streptomyces (Streptomyces) etc., comprising filamentous fungi, yeast and bacterium, existing microbes producing cellulase profitable probliotics again, has possessed abundant and powerful enzymatic productivity, microbe conversion ability and beneficial function, the biology being expected to be applied to lignocellulosic and puffed soybean agricultural by-products transforms.
Edible mushroom is a large class take lignocellulosic as the grade-safe macro fungi that matrix carries out growth and breeding, and important edible mushroom kind comprises flat mushroom (Pleurotus ostreatus), fan-shaped pick up the ears (Pieurotus flabellatus), lung shape picks up the ears (Pleurotus pulmonarius), phoenix-tail mushroom (Pleurotus sajor-caju), mushroom (Lentinus edobes), Lentinus tigrinus (Lentinus tiginus), asparagus (Flamulina velutipes), glossy ganoderma (Gannodema lucidum), schizophyllum commune (Schizophyllum commune), arteries and veins penetrates bacterium (Phlebia radiata), rainbow conk (Coriolus versicolor), the yellow mushroom (Plearotus citrinopileatus) of elm, Ji mushroom (Plearotus cornucopiae), pleurotus eryngii (Pleurotus eryngii), auricularia auriculajudae (Auricularia auricular), hickory chick (Morchella esculenta), agrocybe (Agrocybe cylindracea), yellow umbrella (Pholiota adipose), agaricus bisporus (Agaricus bisporus), the beautiful gill fungus (Hypsizigus marmoreus) of spot, Hericium erinaceus (Hercicium erinaceum), Agricus blazei (Agaricus blazei), sliding mushroom (Pholiota nameko) etc.Because edible mushroom secretes a large amount of lignocellulosic enzyme in growth and breeding process in matrix, thus its matrix to agricultural by-products particularly lignocellulose agricultural by-products there is very strong biotransformation capacity.
Summary of the invention
The invention provides a kind of biology to transform reed and peanut dregs and prepare complex enzyme for feed and to hold concurrently the method for probiotics preparation, adopt ferment reed and peanut dregs of the stable grade-safe flora of Tiny ecosystem to prepare complex enzyme for feed and to hold concurrently probiotics preparation, realize the low-cost high-efficiency biotransformation of reed and peanut dregs.
Biological reed and the peanut dregs of transforming is prepared complex enzyme for feed and to be held concurrently the method for probiotics preparation, comprises the following steps:
1) fully mix through the Pu'er cooked tea of pile-fermentation, lentinus edodes strain stick, agrocybe bacterium rod, the first reed, the first peanut dregs, the first wheat bran, the first pomace and the first water, after domestication is cultivated, form leavening;
2) leavening in step 1), the second reed, the second peanut dregs, the second wheat bran, the second pomace and the second water are fully mixed, after fermentation, obtain complex enzyme for feed and to hold concurrently probiotics preparation.
Pu'er tea is a kind of with a long history, generally regarded as safe drink, the wet heap flora of the Pu'er tea formed in the Pu'er cooked tea of pile-fermentation is grade-safe flora, mushroom and agrocybe are daily edible grade-safe macro fungis, containing abundant lignocellulosic enzyme in its bacterium rod.Reed and peanut dregs are two kinds of large agricultural by-products.By the Pu'er cooked tea through pile-fermentation, lentinus edodes strain stick, agrocybe bacterium rod and the first reed, first peanut dregs, first wheat bran, after the matrix mixing such as the first pomace and the first water, cultivated by domestication, natural flora in Pu'er tea wet heap flora and matrix is merged and forms the stable grade-safe flora of Tiny ecosystem, this flora had both had the abundant and powerful enzymatic productivity of Pu'er tea wet heap flora, microbe conversion ability and beneficial function, accommodate substrate environment again, simultaneously, containing the abundant lignocellulosic enzyme deriving from lentinus edodes strain stick and agrocybe bacterium rod in matrix environment, low-cost high-efficiency biotransformation can be carried out to reed (i.e. the first reed and the second reed) and peanut dregs (i.e. the first peanut dregs and the second peanut dregs).Therefore, the leavening better performances obtained.
The grade-safe flora stable with Tiny ecosystem in leavening is fermented to reed and peanut dregs, reed and peanut dregs is converted into complex enzyme for feed and holds concurrently probiotics preparation, and the hold concurrently property indices of probiotics preparation of the complex enzyme for feed obtained is better.
In leavening preparation with sweat, reed is as primary carbon source, and peanut dregs, as main nitrogen, constitutes the basis in fermentation substrate.Wheat bran (i.e. the first wheat bran and the second wheat bran) is rich in vitamin and mineral element, pomace (i.e. the first pomace and the second pomace) is rich in the fermentable sugar that vitamin and microorganism can utilize fast, adds wheat bran and pomace and can be fermentation process and provide sufficient growth factor (confactor as Some Related Enzymes) and utilize carbon source fast.
In step 1), as preferably, the mass ratio of the described Pu'er cooked tea through pile-fermentation, lentinus edodes strain stick, agrocybe bacterium rod, the first reed, the first peanut dregs, the first wheat bran, the first pomace and the first water is 10:1 ~ 10:1 ~ 10:5 ~ 15:2 ~ 8:1 ~ 4:0.5 ~ 2:10 ~ 30, under the raw material of above-mentioned mass ratio, be conducive to Pu'er tea wet heap flora and natural flora to merge and form the stable grade-safe flora of Tiny ecosystem, obtain the better leavening of fermenting property.Further preferably, the mass ratio of the described Pu'er cooked tea through pile-fermentation, lentinus edodes strain stick, agrocybe bacterium rod, the first reed, the first peanut dregs, the first wheat bran, the first pomace and the first water is 10:5:5:10:5:2:1:22 ~ 25, the leavening made under the raw material of this mass ratio, its fermenting property is the most excellent.
As preferably, the condition that described domestication is cultivated is 30 DEG C ~ 45 DEG C domestication cultivations 24 ~ 60 hours, the condition natural flora be conducive in Pu'er tea wet heap flora and matrix that this domestication is cultivated merges and forms the stable grade-safe flora of Tiny ecosystem, prepares the better leavening of fermenting property.Further preferably, the condition that described domestication is cultivated is that the condition that this domestication is cultivated can prepare the most excellent leavening of fermenting property 35 DEG C ~ 40 DEG C domestication cultivations 36 ~ 48 hours.
Step 2) in, as preferably, the mass ratio of described leavening, the second reed, the second peanut dregs, the second wheat bran, the second pomace and the second water is 10:100 ~ 800:50 ~ 500:10 ~ 100:5 ~ 60:100 ~ 1500, under the raw material of above-mentioned mass ratio, after fermentation, be conducive to obtaining the better complex enzyme for feed of property indices and hold concurrently probiotics preparation.Further preferably, the mass ratio of described leavening, the second reed, the second peanut dregs, the second wheat bran, the second pomace and the second water is 10:300 ~ 600:150 ~ 300:30 ~ 60:15 ~ 30:400 ~ 1000, is conducive to obtaining the best complex enzyme for feed of property indices and holds concurrently probiotics preparation.
As preferably, described fermentation adopts solid state fermentation, and microorganism easily grows, and enzyme activity is high, and preparation is simple, is conducive to industrialization promotion and utilizes.Further preferred, the condition of described solid state fermentation was: material thickness is 5cm ~ 20cm, gravity-flow ventilation, 30 DEG C ~ 45 DEG C fermentations 2 ~ 7 days.This solid state fermentation conditions is conducive to obtaining the more excellent complex enzyme for feed of property indices holds concurrently probiotics preparation.Further preferably, the condition of described solid state fermentation is: the material thickness of solid state fermentation is 10cm ~ 15cm, gravity-flow ventilation, 35 DEG C ~ 40 DEG C fermentations 4 ~ 5 days, this solid state fermentation conditions is conducive to obtaining the most excellent complex enzyme for feed of property indices held concurrently probiotics preparation.
Nutritional labeling in feed divides the mainly compound (as lignin, alpha-glucosides class, phytic acid, tannin etc.) such as carbohydrate (as starch, cellulose, hemicellulose, pectic substance), protein, heterocyclic and atypia sugar according to chemical composition.In feed, add the enzyme these compounds being carried out to predigestion or conversion, for raising livestock and poultry production performance, there is active effects.At present, cellulase, hemicellulase such as zytase, amylase, protease, alpha-galactosidase, phytase etc. all widely use in Feed Manufacturing as enzyme Preparations Used for Feeds, and in use usually the multiple enzyme of combination become complex enzyme formulation and add.The enzyme comprised in complex enzyme formulation is more, and the effect of feed being carried out to predigestion or conversion is higher.The flora that the present invention adopts is that the natural flora in Pu'er tea wet heap flora and fermentation substrate merges the stable grade-safe flora of the Tiny ecosystem that formed, microbe species enriches, fermenting and producing can go out the complex enzyme for feed that kind enriches very much, to the evaluation of its effect by carrying out the mensuration of each Some Related Enzymes vigor.
Probio has remarkable efficacy by conditioning function of intestinal canal to lifting animal health level.Lactic acid bacteria is topmost probio.In the flora that the present invention adopts, topmost source is the wet heap flora of Pu'er tea, and wherein containing a large amount of lactic acid bacteria, these lactic acid bacterias breed in a large number through solid state fermentation, become probiotics preparation, to the evaluation of its effect by carrying out the mensuration of lactic acid bacterium number.
By each measurement result of Some Related Enzymes vigor and the measurement result of lactic acid bacterium number, known, product prepared by the present invention is suitable as complex enzyme for feed and holds concurrently probiotics preparation very much.
Compared with prior art, tool of the present invention has the following advantages:
In the present invention, biological reed and the peanut dregs of transforming is prepared complex enzyme for feed and to be held concurrently the method for probiotics preparation, by the Pu'er cooked tea through pile-fermentation, lentinus edodes strain stick, agrocybe bacterium rod and the first reed, first peanut dregs, first wheat bran, after the matrix mixing such as the first pomace and the first water, cultivated by domestication, Pu'er tea wet heap flora in the Pu'er cooked tea of pile-fermentation and the natural flora in matrix are merged and forms the stable grade-safe flora of Tiny ecosystem, the stable grade-safe flora of this Tiny ecosystem had both had the abundant and powerful enzymatic productivity of Pu'er tea wet heap flora, microbe conversion ability and beneficial function, accommodate substrate environment again, simultaneously, containing the abundant lignocellulosic enzyme deriving from lentinus edodes strain stick and agrocybe bacterium rod in matrix environment, low-cost high-efficiency biotransformation can be carried out to reed and peanut dregs.The grade-safe flora stable with Tiny ecosystem in leavening is fermented to reed and peanut dregs, reed and peanut dregs is converted into complex enzyme for feed and holds concurrently probiotics preparation, and the hold concurrently property indices of probiotics preparation of the complex enzyme for feed obtained is better.
In the present invention, biological reed and the peanut dregs of transforming is prepared complex enzyme for feed and to be held concurrently the method for probiotics preparation, under the grade-safe flora effect that Tiny ecosystem is stable in leavening, fermentation can adopt solid state fermentation, microorganism easily grows, and enzyme activity is high, and sweat is extensive, do not need stringent asepsis requirements, post processing is easy, non-wastewater discharge, and simple in equipment, small investment, energy consumption are low, easy to operate, be easy to industrialization large-scale production, there is good economic benefit and wide application prospect.
Detailed description of the invention
The commercially available prod that lentinus edodes strain stick in embodiment and agrocybe bacterium rod all adopt the Tianquan Moganshan Mountain, Zhejiang Gu Ye Co., Ltd to produce, reed gathers voluntarily from planting site, peanut dregs adopts the commercially available prod of Queshan County Sanlihe Zhang Shi grouts Trade Department, the commercially available prod that wheat bran adopts Jin Maiyuanmai benevolence factory of Dafeng City to produce, the commercially available prod that pomace adopts the two flourish ecological Farming Ltd. in Wanrong County to produce.
Embodiment 1
1) 10kg is fully mixed with 10kg reed, 5kg peanut dregs, 2kg wheat bran, 1kg pomace, 22kg water through the Pu'er cooked tea of pile-fermentation, 5kg lentinus edodes strain stick, 5kg agrocybe bacterium rod, 35 DEG C of domestication cultivations after 48 hours, formation leavening;
2) the leavening 20kg in step 1) is fully mixed with 600kg reed, 300kg peanut dregs, 60kg wheat bran, 30kg pomace, 800kg water, adopt solid state fermentation, material thickness is 15cm, gravity-flow ventilation, 40 DEG C of fermentations 4 days, obtain complex enzyme for feed and to hold concurrently probiotics preparation.
After measured, the hold concurrently performance indications of probiotics preparation of the complex enzyme for feed of the present embodiment are: carboxymethylcelluloenzyme enzyme activity reaches 48.58IU/g; Total fiber element enzyme activity reaches 4.87IU/g; Beta-glucoside enzyme activity reaches 12.12IU/g; Endoglucanase enzyme activity reaches 39.85IU/g; Xylanase activity reaches 41.51IU/g; Beta-xylosidase vigor reaches 3.72IU/g; Polygalacturonase vigor reaches 77.55IU/g; Pectin lyase vigor reaches 74.46IU/g; Laccase activity reaches 87.26IU/g; Mannosan enzyme activity reaches 35.38IU/g; Alpha-galactoside enzyme activity reaches 10.20IU/g; Alpha amylase activity reaches 140.06IU/g; Prolease activity reaches 124637IU/g; Phytase activity reaches 110.50IU/g; Tannase vigor reaches 1327IU/g; Lactic acid bacteria density reaches 2.82 × 10
8cFU/g.
Embodiment 2
1) 10kg is fully mixed with 10kg reed, 5kg peanut dregs, 2kg wheat bran, 1kg pomace, 25kg water through the Pu'er cooked tea of pile-fermentation, 5kg lentinus edodes strain stick, 5kg agrocybe bacterium rod, 35 DEG C of domestication cultivations after 48 hours, formation leavening;
2) the leavening 20kg in step 1) is fully mixed with 600kg reed, 300kg peanut dregs, 60kg wheat bran, 30kg pomace, 800kg water, adopt solid state fermentation, material thickness is 15cm, gravity-flow ventilation, 40 DEG C of fermentations 4 days, obtain complex enzyme for feed and to hold concurrently probiotics preparation.
After measured, the hold concurrently performance indications of probiotics preparation of the complex enzyme for feed of the present embodiment are: carboxymethylcelluloenzyme enzyme activity reaches 48.19IU/g; Total fiber element enzyme activity reaches 4.96IU/g; Beta-glucoside enzyme activity reaches 12.08IU/g; Endoglucanase enzyme activity reaches 39.64IU/g; Xylanase activity reaches 41.48IU/g; Beta-xylosidase vigor reaches 3.68IU/g; Polygalacturonase vigor reaches 77.35IU/g; Pectin lyase vigor reaches 74.32IU/g; Laccase activity reaches 87.18IU/g; Mannosan enzyme activity reaches 35.87IU/g; Alpha-galactoside enzyme activity reaches 10.12IU/g; Alpha amylase activity reaches 140.14IU/g; Prolease activity reaches 124768IU/g; Phytase activity reaches 108.56IU/g; Tannase vigor reaches 1325IU/g; Lactic acid bacteria density reaches 2.80 × 10
8cFU/g.
Embodiment 3
1) 10kg is fully mixed with 10kg reed, 5kg peanut dregs, 2kg wheat bran, 1kg pomace, 25kg water through the Pu'er cooked tea of pile-fermentation, 5kg lentinus edodes strain stick, 5kg agrocybe bacterium rod, 40 DEG C of domestication cultivations after 36 hours, formation leavening;
2) the leavening 10kg in step 1) is fully mixed with 600kg reed, 300kg peanut dregs, 60kg wheat bran, 30kg pomace, 1000kg water, adopt solid state fermentation, material thickness is 10cm, gravity-flow ventilation, 35 DEG C of fermentations 5 days, obtain complex enzyme for feed and to hold concurrently probiotics preparation.
After measured, the hold concurrently performance indications of probiotics preparation of the complex enzyme for feed of the present embodiment are: carboxymethylcelluloenzyme enzyme activity reaches 47.70IU/g; Total fiber element enzyme activity reaches 5.22IU/g; Beta-glucoside enzyme activity reaches 11.84IU/g; Endoglucanase enzyme activity reaches 41.12IU/g; Xylanase activity reaches 40.80IU/g; Beta-xylosidase vigor reaches 3.60IU/g; Polygalacturonase vigor reaches 76.41IU/g; Pectin lyase vigor reaches 72.44IU/g; Laccase activity reaches 86.46IU/g; Mannosan enzyme activity reaches 33.07IU/g; Alpha-galactoside enzyme activity reaches 9.90IU/g; Alpha amylase activity reaches 134.15IU/g; Prolease activity reaches 130212IU/g; Phytase activity reaches 112.43IU/g; Tannase vigor reaches 1159IU/g; Lactic acid bacteria density reaches 2.86 × 10
8cFU/g.
Embodiment 4
1) 10kg is fully mixed with 5kg reed, 2kg peanut dregs, 1kg wheat bran, 0.5kg pomace, 10kg water through the Pu'er cooked tea of pile-fermentation, 1kg lentinus edodes strain stick, 1kg agrocybe bacterium rod, 45 DEG C of domestication cultivations after 24 hours, formation leavening;
2) the leavening 20kg in step 1) is fully mixed with 200kg reed, 100kg peanut dregs, 20kg wheat bran, 10kg pomace, 200kg water, adopt solid state fermentation, material thickness is 5cm, gravity-flow ventilation, 45 DEG C of fermentations 2 days, obtain complex enzyme for feed and to hold concurrently probiotics preparation.
After measured, the hold concurrently performance indications of probiotics preparation of the complex enzyme for feed of the present embodiment are: carboxymethylcelluloenzyme enzyme activity reaches 43.45IU/g; Total fiber element enzyme activity reaches 4.74IU/g; Beta-glucoside enzyme activity reaches 10.78IU/g; Endoglucanase enzyme activity reaches 37.43IU/g; Xylanase activity reaches 36.68IU/g; Beta-xylosidase vigor reaches 3.32IU/g; Polygalacturonase vigor reaches 70.11IU/g; Pectin lyase vigor reaches 75.32IU/g; Laccase activity reaches 78.34IU/g; Mannosan enzyme activity reaches 30.62IU/g; Alpha-galactoside enzyme activity reaches 9.12IU/g; Alpha amylase activity reaches 122.46IU/g; Prolease activity reaches 120325IU/g; Phytase activity reaches 104.56IU/g; Tannase vigor reaches 1027IU/g; Lactic acid bacteria density reaches 2.53 × 10
8cFU/g.
Embodiment 5
1) 10kg is fully mixed with 15kg reed, 8kg peanut dregs, 4kg wheat bran, 2kg pomace, 30kg water through the Pu'er cooked tea of pile-fermentation, 10kg lentinus edodes strain stick, 10kg agrocybe bacterium rod, 30 DEG C of domestication cultivations after 60 hours, formation leavening;
2) the leavening 10kg in step 1) is fully mixed with 800kg reed, 500kg peanut dregs, 100kg wheat bran, 60kg pomace, 1500kg water, adopt solid state fermentation, material thickness is 20cm, gravity-flow ventilation, 30 DEG C of fermentations 7 days, obtain complex enzyme for feed and to hold concurrently probiotics preparation.
After measured, the hold concurrently performance indications of probiotics preparation of the complex enzyme for feed of the present embodiment are: carboxymethylcelluloenzyme enzyme activity reaches 43.34IU/g; Total fiber element enzyme activity reaches 4.65IU/g; Beta-glucoside enzyme activity reaches 10.59IU/g; Endoglucanase enzyme activity reaches 37.34IU/g; Xylanase activity reaches 36.45IU/g; Beta-xylosidase vigor reaches 3.34IU/g; Polygalacturonase vigor reaches 69.87IU/g; Pectin lyase vigor reaches 75.87IU/g; Laccase activity reaches 77.68IU/g; Mannosan enzyme activity reaches 30.48IU/g; Alpha-galactoside enzyme activity reaches 9.06IU/g; Alpha amylase activity reaches 122.64IU/g; Prolease activity reaches 120435IU/g; Phytase activity reaches 105.23IU/g; Tannase vigor reaches 1034IU/g; Lactic acid bacteria density reaches 2.51 × 10
8cFU/g.
Various enzyme activity and lactic acid bacteria density inspect method as follows:
One, the mensuration of the enzyme activity of carboxymethylcelluloenzyme enzyme, total fiber element enzyme and beta-glucuroide
Total fiber element enzyme (namely carry out (referring to " Ghose TK; 1987.Measurements of cellulase activities.Pure Appl Chem; 59,257-268. ") according to the established procedure of international pure chemistry and applied chemistry federation (International Union of Pure and Applied Chemistry) by the mensuration of Filter paper Cellulase (FPase), carboxymethylcelluloenzyme enzyme (CMCase), GBAP vigor.
Filter paper Cellulase (FPase) unit of activity: with Whatman No.1 filter paper (1.0 × 6.0cm=50.0mg) for substrate, at 50 DEG C, reaction 60min in 50mM sodium citrate buffer solution (pH=4.8), be an enzyme activity international unit (IU) with release per minute 1 μm of ol reduced sugar, be expressed as IU/g dry weight.
Carboxymethylcelluloenzyme enzyme (CMCase) unit of activity: with 2% (w/v) carboxymethyl cellulose for substrate, at 50 DEG C, reaction 30min in 50mM sodium citrate buffer solution (pH=5.5), be an enzyme activity international unit (IU) with release per minute 1 μm of ol reduced sugar, be expressed as IU/g dry weight.
GBAP unit of activity: add 1mL paranitrophenol-beta-glucosidase (p-nitrophenyl glucopyranoside in 2mL acetate buffer, pNPG) (1mM) is as substrate, 30min is reacted at 50 DEG C, 430nm colorimetric estimation, be an enzyme activity international unit (IU) with release per minute 1 μm of ol paranitrophenol (p-nitrophenol) (pNP), be expressed as IU/g dry weight.
Two, the mensuration of the enzyme activity of endoglucanase, zytase, beta-xylosidase, polygalacturonase and Pectin lyase
Endoglucanase, zytase, polygalacturonase and Pectin lyase vitality test are respectively with carboxymethyl cellulose, birch xylan enzyme, polygalacturonic acid and pectic acid are substrate, specifically according to related documents (Cheilas T, Stoupis T, Christakopoulos P, Katapodis P, Mamma D, Hatzinikolaou DG, Kekos D, Macris BJ, 2000.Hemicellulolytic activity of Fusarium oxysporum grown on sugar beet pulp.Production of extracellular arabinanase.Process Biochem, 35, 557-561, Jayani RS, Saxena S, Gupta R, 2005.Microbial pectinolytic enzymes:a review.Process Biochem, 40,2931-2944.).Release per minute 1 μm of ol reduced sugar is an enzyme activity international unit (IU), is expressed as IU/g dry weight.
Beta-xylosidase vitality test with nitre phenyl glycoside (p-nitrophenyl glycoside) for substrate, specifically according to related documents (Mamma D, Koullas D, Fountoukidis G, Kekos D, Macris BJ, Koukios E, 1996.Bioethanol from sweet sorghum:Simultaneous saccharification and fermentation of carbohydrates by a mixed microbial culture.Process Biochem, 31,377-381.).Release per minute 1 μm of ol p-nitrophenol is an enzyme activity international unit (IU), is expressed as IU/g dry weight.
Three, the mensuration of the enzyme activity of laccase
Laccase activity passes through ABTS (2, 2 '-azino-bis-3-ethylbenzthiazoline-6-sulfonic acid) oxidation and measure, reaction system is that the 20mM ABTS adding 100 μ L in sodium citrate buffer solution (pH=3.0) uses sodium citrate buffer solution (pH=3.0) to be settled to 1mL again, ABTS oxygenation efficiency is determined by 420nm absorption value, specifically according to related documents (Susan Grace Karp, Vincenza Faraco, Antonella Amore, Leila Birolo, Chiara Giangrande, Vanete Thomaz Soccol, Ashok Pandey, Carlos Ricardo Soccol.Characterization of laccase isoforms produced by Pleurotus ostreatus in solid state fermentation of sugarcane bagasse.Bioresource Technology, 2012, 114, 735-739.).Oxidation per minute 1 μm of ol ABTS is an enzyme activity international unit (IU), is expressed as IU/g dry weight.
Four, the mensuration of the enzyme activity of mannase
The vitality test of mannase take carob as substrate, 1g enzyme powder in 40 DEG C, under pH5.0 condition, 1min is hydrolyzed carob and generates and be equivalent to 1 μm of ol mannose reducing substances, is 1 enzyme activity unit, is expressed as IU/g dry weight.
Five, the mensuration of the enzyme activity of alpha-galactosidase
Alpha-galactoside enzyme activity determination system is " 0.1mL enzyme liquid+0.8mL0.2M acetate buffer (pH4.8)+0.1mL2mM pNPG ", after 50 DEG C of reaction 15min, add 3mL0.2MNa2CO3 cessation reaction, 405nm place surveys absorption value, specifically according to related documents (Dey PM, Pridham JB, 1972.Biochemistry of alpha-galactosidase.Adv Enzymol, 36,911-930.).Release per minute 1 μm of ol paranitrophenol (paranitrophenol) is an enzyme activity international unit (IU), is expressed as IU/g dry weight.
Six, the mensuration of the enzyme activity of AMY
AMY vitality test system is " 150 μ L enzyme liquid+200 μ L0.2% soluble starches, with 0.1M Tris-HCl buffer solution (pH=7.0) for solution system ", after 37 DEG C of reaction 30min, add 400 μ l3, 5-dinitro salicylic acid cessation reaction boiling water bath keep 5min, 8mL distilled water diluting is added after being cooled to room temperature 25 DEG C, 489nm place surveys absorption value, specifically according to related documents (Bernfeld P (1955) Amylases, alpha and beta.In:Colowick SP, Kaplan NO (eds) Methods in enzymology, Vol1.Academic, New York, pp149-154.).Release per minute 1 μm of ol reduced sugar is an enzyme activity international unit (IU), is expressed as IU/g dry weight.
Seven, the mensuration of the enzyme activity of protease
Prolease activity measures with sulphanilamide azo-casein (sulphanilamide azocasein) as substrate, reaction system is containing 0.5% azo-casein (azocasein) (w/v) in 250 μ L0.1M phosphate buffers (pH8.5), add 150 μ L enzyme liquid again, after 37 DEG C of reaction 30min, add 1.2mL solution of trichloroacetic acid (trichloroacetic acid solution) (10%, w/v) go out enzyme, add 800 μ L of1.8N NaOH again to neutralize, 420nm place surveys absorption value, specifically according to related documents (Leighton TJ, Doi RH, Warren RAJ, Kelln RA (1973) The relationship of serine protease activity to RNA polymerase modification and sporulation in Bacillus subtilis.J Mol Biol, 76:103-122.). release 1 μ g azo-casein (azocasein) per minute is an enzyme activity international unit (IU), be expressed as IU/g dry weight.
Eight, the mensuration of the enzyme activity of phytase and tannase
Phytase activity measures with para-nitro-pheneye phosphate (p-nitrophenylphosphate) as substrate, specifically according to related documents (Stockmann C, Losen M, Dahlems U, Knocke C, Gellissen G, Buchs J (2003) .Effect of oxygen supply on passaging, stabilizing and screening of recombinant Hansenula polymorpha production strains in test tube cultures.FEMS Yeast Res, 4 (2): 195-205.).Release per minute 1 μm of ol p-nitrophenol (p-nitrophenol) is an enzyme activity international unit (IU), is expressed as IU/g dry weight.
Tannase vitality test take tannic acid as substrate, specifically according to related documents (Mondal KC, Banerjee D, Jana M, Pati BR (2001) .Colorimetric assay method for determination of the tannin acyl hydrolase (EC3.1.1.20) activity.Anal Biochem, 295 (2): 168-171.).Conversion per minute 1 μm of ol tannic acid is an enzyme activity international unit (IU), is expressed as IU/g dry weight.
Nine, the mensuration of lactic acid bacteria density
Man Rogosa Sharpe (MRS) culture medium is lactic acid bacteria Selective agar medium, measures the total plate count on Man Rogosa Sharpe (MRS) culture medium, can be scaled lactic acid bacteria density.Total plate count measures carries out (Song Y according to related documents, Luo Y, You J, Shen H, & Hu S. (2012) .Biochemical, sensory and microbiological attributes of bream (Megalobrama amblycephala) during partial freezing and chilled storage.J Sci Food Agric, 92 (1), 197-202.).
Claims (5)
1. biological reed and the peanut dregs of transforming is prepared complex enzyme for feed and to be held concurrently the method for probiotics preparation, it is characterized in that, comprises the following steps:
1) fully mix through the Pu'er cooked tea of pile-fermentation, lentinus edodes strain stick, agrocybe bacterium rod, the first reed, the first peanut dregs, the first wheat bran, the first pomace and the first water, after domestication is cultivated, form leavening;
The mass ratio of the described Pu'er cooked tea through pile-fermentation, lentinus edodes strain stick, agrocybe bacterium rod, the first reed, the first peanut dregs, the first wheat bran, the first pomace and the first water is 10:1 ~ 10:1 ~ 10:5 ~ 15:2 ~ 8:1 ~ 4:0.5 ~ 2:10 ~ 30;
The condition that described domestication is cultivated is 30 DEG C ~ 45 DEG C domestication cultivations 24 ~ 60 hours;
2) by step 1) in leavening, the second reed, the second peanut dregs, the second wheat bran, the second pomace and the second water fully mix, after fermentation, obtain complex enzyme for feed and to hold concurrently probiotics preparation;
The mass ratio of described leavening, the second reed, the second peanut dregs, the second wheat bran, the second pomace and the second water is 10:100 ~ 800:50 ~ 500:10 ~ 100:5 ~ 60:100 ~ 1500;
Described fermentation adopted solid state fermentation, and the condition of described solid state fermentation is: material thickness is 5cm ~ 20cm, gravity-flow ventilation, 30 DEG C ~ 45 DEG C fermentations 2 ~ 7 days.
2. biology according to claim 1 transforms reed and peanut dregs and prepares complex enzyme for feed and to hold concurrently the method for probiotics preparation, it is characterized in that, step 1) in, the mass ratio of the described Pu'er cooked tea through pile-fermentation, lentinus edodes strain stick, agrocybe bacterium rod, the first reed, the first peanut dregs, the first wheat bran, the first pomace and the first water is 10:5:5:10:5:2:1:22 ~ 25.
3. biology according to claim 1 transforms reed and peanut dregs and prepares complex enzyme for feed and to hold concurrently the method for probiotics preparation, it is characterized in that, step 1) in, the condition that described domestication is cultivated be tame cultivations 36 ~ 48 hours at 35 DEG C ~ 40 DEG C.
4. biology according to claim 1 transforms reed and peanut dregs and prepares complex enzyme for feed and to hold concurrently the method for probiotics preparation, it is characterized in that, step 2) in, the mass ratio of described leavening, the second reed, the second peanut dregs, the second wheat bran, the second pomace and the second water is 10:300 ~ 600:150 ~ 300:30 ~ 60:15 ~ 30:400 ~ 1000.
5. biology according to claim 1 transforms reed and peanut dregs and prepares complex enzyme for feed and to hold concurrently the method for probiotics preparation, it is characterized in that, step 2) in, the condition of described solid state fermentation is: the material thickness of solid state fermentation is 10cm ~ 15cm, gravity-flow ventilation, 35 DEG C ~ 40 DEG C fermentations 4 ~ 5 days.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201310229627.XA CN103300210B (en) | 2013-06-08 | 2013-06-08 | Method for preparing compound enzyme and probiotic preparation for feed by biotransformation of reed and peanut cake |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201310229627.XA CN103300210B (en) | 2013-06-08 | 2013-06-08 | Method for preparing compound enzyme and probiotic preparation for feed by biotransformation of reed and peanut cake |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN103300210A CN103300210A (en) | 2013-09-18 |
| CN103300210B true CN103300210B (en) | 2015-01-07 |
Family
ID=49126277
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201310229627.XA Expired - Fee Related CN103300210B (en) | 2013-06-08 | 2013-06-08 | Method for preparing compound enzyme and probiotic preparation for feed by biotransformation of reed and peanut cake |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN103300210B (en) |
Families Citing this family (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105124138A (en) * | 2015-10-12 | 2015-12-09 | 河北大学 | High-protein reed feed and preparation method thereof |
| CN108497179A (en) * | 2018-04-17 | 2018-09-07 | 湖南诚扬生物科技有限公司 | A kind of high protein ensiling reed feed and preparation method improving beef quality |
Citations (14)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1041264A (en) * | 1988-08-30 | 1990-04-18 | 池田久和 | Edible material |
| CN1049958A (en) * | 1990-09-12 | 1991-03-20 | 李庆义 | The method of processing feedstuff with stalks of crops |
| CN1579197A (en) * | 2004-05-18 | 2005-02-16 | 王升厚 | Bacterisugar mono-cell protein feed producing method |
| CN1653936A (en) * | 2005-04-05 | 2005-08-17 | 迟乃玉 | Method for preparing composite micro-ecological enzyme for feeding the ruminant by using disused mushroom dregs |
| CN1711885A (en) * | 2004-06-24 | 2005-12-28 | 王冬 | Non-pollution nuisance composite mycelium active forage and production thereof |
| CN1934985A (en) * | 2006-10-24 | 2007-03-28 | 南京师范大学 | Method for producing fodder by in-situ post fermentation of flammulina velutipes bran mycelia |
| CN101124937A (en) * | 2007-09-07 | 2008-02-20 | 梁亚桐 | Technology for producing biological fermentation complete feed |
| CN101584466A (en) * | 2008-05-22 | 2009-11-25 | 刘泳宏 | Natural lung-nourishing asthma-relieving egg and preparation method thereof |
| CN101692864A (en) * | 2009-10-16 | 2010-04-14 | 胡宁华 | Tea organism mixed feed and preparation method thereof |
| CN101812431A (en) * | 2009-02-23 | 2010-08-25 | 湖南省畜牧兽医研究所 | Method utilizing bacterisugar to prepare feed enzyme |
| CN101822311A (en) * | 2010-05-07 | 2010-09-08 | 吴建敏 | Process for producing solid biologic ferment by utilizing biogas residue |
| CN102715339A (en) * | 2012-06-01 | 2012-10-10 | 陈华友 | Microorganism fodder production method based on pleurotus eryngii mushroom cultivating residues |
| CN103070308A (en) * | 2013-01-09 | 2013-05-01 | 李峰 | Feed for nursing rabbits and preparation method thereof |
| CN103082145A (en) * | 2013-01-29 | 2013-05-08 | 大连工业大学 | Method for producing grape skin residue pig feed by utilizing lentinula edodes and yeast for symbiotic fermentation |
-
2013
- 2013-06-08 CN CN201310229627.XA patent/CN103300210B/en not_active Expired - Fee Related
Patent Citations (14)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1041264A (en) * | 1988-08-30 | 1990-04-18 | 池田久和 | Edible material |
| CN1049958A (en) * | 1990-09-12 | 1991-03-20 | 李庆义 | The method of processing feedstuff with stalks of crops |
| CN1579197A (en) * | 2004-05-18 | 2005-02-16 | 王升厚 | Bacterisugar mono-cell protein feed producing method |
| CN1711885A (en) * | 2004-06-24 | 2005-12-28 | 王冬 | Non-pollution nuisance composite mycelium active forage and production thereof |
| CN1653936A (en) * | 2005-04-05 | 2005-08-17 | 迟乃玉 | Method for preparing composite micro-ecological enzyme for feeding the ruminant by using disused mushroom dregs |
| CN1934985A (en) * | 2006-10-24 | 2007-03-28 | 南京师范大学 | Method for producing fodder by in-situ post fermentation of flammulina velutipes bran mycelia |
| CN101124937A (en) * | 2007-09-07 | 2008-02-20 | 梁亚桐 | Technology for producing biological fermentation complete feed |
| CN101584466A (en) * | 2008-05-22 | 2009-11-25 | 刘泳宏 | Natural lung-nourishing asthma-relieving egg and preparation method thereof |
| CN101812431A (en) * | 2009-02-23 | 2010-08-25 | 湖南省畜牧兽医研究所 | Method utilizing bacterisugar to prepare feed enzyme |
| CN101692864A (en) * | 2009-10-16 | 2010-04-14 | 胡宁华 | Tea organism mixed feed and preparation method thereof |
| CN101822311A (en) * | 2010-05-07 | 2010-09-08 | 吴建敏 | Process for producing solid biologic ferment by utilizing biogas residue |
| CN102715339A (en) * | 2012-06-01 | 2012-10-10 | 陈华友 | Microorganism fodder production method based on pleurotus eryngii mushroom cultivating residues |
| CN103070308A (en) * | 2013-01-09 | 2013-05-01 | 李峰 | Feed for nursing rabbits and preparation method thereof |
| CN103082145A (en) * | 2013-01-29 | 2013-05-08 | 大连工业大学 | Method for producing grape skin residue pig feed by utilizing lentinula edodes and yeast for symbiotic fermentation |
Non-Patent Citations (1)
| Title |
|---|
| 茶叶微生物研究进展及展望;胥伟 等;《福建茶叶》;20090625(第2期);第6页右栏第2-3段，第7页右栏倒数第2段"1.5 茶叶废弃物与微生物" * |
Also Published As
| Publication number | Publication date |
|---|---|
| CN103300210A (en) | 2013-09-18 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN103355477B (en) | Production method for feed through fermentation of soy sauce residues | |
| CN103300213B (en) | Method for preparing compound enzyme and probiotic preparation for feed through biotransformation of broad bean stems/leaves and cottonseed meals | |
| CN103315131B (en) | Preparation method of compound enzymes and probiotics for forage by biotransformation of broccoli rhizome and soybean cake | |
| Liu et al. | Solid-state fermentation of ammoniated corn straw to animal feed by Pleurotus ostreatus Pl-5 | |
| CN103749987A (en) | Coarse cereal, mixed meal and daily ration enzyme preparation and preparation method thereof | |
| CN103300210B (en) | Method for preparing compound enzyme and probiotic preparation for feed by biotransformation of reed and peanut cake | |
| CN103340281B (en) | Preparation method of compound enzyme and probiotic preparation for feed through biotransformation of potato stem leaf and sunflower cake | |
| CN103340279B (en) | Method of using biotransformation of rice straws and peanut cakes to prepare composite enzyme and probiotics preparation for forage | |
| CN103300212B (en) | Method for preparing compound enzyme and probiotic preparation for feed through biotransformation of corn stalks and soybean meals | |
| CN103283947B (en) | Method for preparing complex enzyme-probiotic preparation for feeds through biotransformation of barley straws and tea seed cakes | |
| CN103300215B (en) | Method for preparing compound enzyme and probiotic preparation for feed through biotransformation of pea stems/leaves and palm meals | |
| CN103315137B (en) | Method for preparing composite enzyme/probiotic preparation for feed by bioconverting rape stem and leaf and rapeseed cake pulp | |
| CN103300214B (en) | Method for preparing compound enzyme and probiotic preparation for feed through biotransformation of corn stalks and cottonseed meals | |
| CN103283945B (en) | Method for preparing complex enzyme-probiotic preparation for feeds through biotransformation of bagasse and sunflower cakes | |
| CN103315133B (en) | Method for preparing composite enzyme/probiotic preparation for feed by bioconverting watermelon vine and olive cake pulp | |
| CN103315134B (en) | Method for preparing composite enzyme/probiotic preparation for feed by bioconverting pumpkin vine and leaf and olive cake pulp | |
| Kahil et al. | Economic co-production of cellulase and αamylase by fungi grown on agro-industrial wastes using solid-state fermentation conditions | |
| CN103283966B (en) | Method for preparing complex enzyme-probiotic preparation for feeds through biotransformation of wheat straws and palm cakes | |
| CN103315135B (en) | Method for preparing composite enzyme/probiotic preparation for feed by bioconverting sweet potato stem and leaf and sesame cake pulp | |
| CN103283946B (en) | Method for preparing complex enzyme-probiotic preparation for feeds through biotransformation of corncobs and sesame seed cakes | |
| CN103315136B (en) | Method for preparing composite enzyme/probiotic preparation for feed by bioconverting soybean stem and leaf and tea seed cake pulp | |
| CN103315132B (en) | Method for preparing composite enzyme/probiotic preparation for feed by bioconverting rice straw and castor cake pulp | |
| CN103300211B (en) | Method for preparing compound enzyme and probiotic preparation for feed by biotransformation of luffa stem and castor cake | |
| CN103300216B (en) | Method for preparing compound enzyme and probiotic preparation for feed through biotransformation of sorghum straws and rapeseed meals | |
| CN103340280B (en) | Method of using biotransformation of corncobs and rapeseed cakes to prepare composite enzyme and probiotics preparation for forage |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C14 | Grant of patent or utility model | ||
| GR01 | Patent grant | ||
| CF01 | Termination of patent right due to non-payment of annual fee | ||
| CF01 | Termination of patent right due to non-payment of annual fee |
Granted publication date: 20150107 Termination date: 20190608 |