The process of solid fermentation of pear slag culture dating polyvispora spores.

Abstract: In order to make full use of pear slag as a resource, the paper takes fresh pear slag as the main raw material, uses solid fermentation method, studies the proportion of pear slag and bran, nitrogen source, inorganic salt, bottling volume, culture temperature and time on the growth and production of spores with the use of polyvixsssss.
research has found that pear slag is a good solid fermentation raw material, suitable for the cultivation of polyvixasporine bacteria.
the maximum number of living cells is obtained when the mass ratio of pear residue to bran is 5:2, 2.5 x 109 cfu/g.
adding urea, sodium nitrate and bean cake powder to the pear slag fermentation medium can significantly improve the number of active bacteria and spores, among which the effect of adding bean cake powder is better. the addition of a mass fraction of 0.5% dihydrophosphate to the
medium media greatly contributes to bacterial growth and the formation of spores.
as the amount of medium bottling increases, the number of spores decreases significantly.
the more suitable culture temperature of polyviable Bacillus D1 is 34 degrees C and the more suitable culture time is 120 h.
the number of live bacteria, spores and spores under suitable culture conditions were 5.09 x 109 cfu/g, 4.78 x 109 cfu/g and 93.9%, respectively.
the results of this study provide a reference for the production of polyviscosine Spore bacteria by fermentation of pear slag.
Introduction: With the increasing area and yield of pear fruit in China, the deep processing of pears has become one of the important ways to increase value after harvesting.
pear juice is one of the main products of pear fruit processing.
produces a large amount of pear slag during the processing of pear juice, the total amount of which is about 40% to 50% of the fruit quality.
, for example, the 10,000 t-natural pear juice factory, which throws away 2,000 to 3,000 t of pear residue per year.
pear residue there are a large number of stone cells, directly used as feed taste is poor, at present basically as waste discarded, not only pollutes the environment, but also causes waste of resources.
if the development and utilization of pear slag can not only increase the economic benefits of enterprises, but also reduce the pollution caused by the accumulation of pear slag to the environment.
Paenibacillus polymyxa is a Grameen-positive bacteria of Bacillaceae-like Bacansporae, also known as polyspora spores, which are not pathogenic to humans or plants.
multi-stick spore bacteria is a plant root probiotic bacteria, can produce peptide antibiotics, antagonistic proteins, enzymes, plant hormones and other physiological living substances, on the one hand can be used as microbial fertilizer, promote plant growth, improve crop yield; The prevention and control of plant bacterial and fungal soil diseases, plant hymn disease, tomato withering disease, rape rot disease, black pine root rot disease and other plant diseases have good prevention and control effect, but also to prevent and control a variety of plant diseases caused by egg bacteria, nematodes.
, polystick spores are a kind of promising and application value-resistant bacteria, which has been used in the production of bio-organic fertilizer, microbial fertilizer and biopesticides.
currently marketed the agent's dosage form is mainly fine granules and wetable powders.
spores are circular or elliptical, thick walls, low water content and strong resistance to the dormant structure formed by spores in cells later in growth and development.
spores have many kinds of resistance, such as heat, drought, ultraviolet light and organic solvents, which is conducive to the survival of Bacillus spores in the environment, the processing and transportation of its bacteriological agents, and is an ideal form for the preparation of microbial preparations.
liquid deep fermentation is widely used in the production of Bacillus spores preparation method.
compared with the following, solid fermentation has the following advantages: the medium is widely sourced and low in price, the equipment used is relatively simple, the energy consumption is low, the production rate of the product is higher, the fermentation process does not discharge waste water, the environmental pollution is less, and the production cost is much lower than the deep fermentation of liquid.
research on polyspora spores culture has focused on liquid fermentation, and no reports of solid fermentation have been seen.
this study, with pear slag as the main raw material, using solid fermentation, the influence of culture conditions on the growth and production of spores d1 of polystick spores, with a view to providing reference for the production of polystick spores with the use of pear slag fermentation.
1 Materials and Methods 1.1 Main Ingredients Fresh Pear Slag: Provided by Zhao Xian Xuhai Juice Co., Ltd., it was measured with a moisture content of 76.2%, a residual sugar quality score of 5.5% and a protein score of 0.32%.
bran: produced at Yongfeng Flour Mill in Yancheng, the water content was 13.2%, the starch mass score was 48.7%, and the protein score was 15.3%.
1.1.2 P.polyvixxa (P.polymyxa)D1: provided by Shijiazhuang Volkswagen Fertilizer Co., Ltd.'s Research and Development Center for Agricultural Microbiobacteria.
1.1.3 Media Bevel Medium: Protein 10.0 g/L, sodium chloride 10.0 g/L, enzyme mother dip powder 5.0 g/L, pH 7.0 (before sterilization).
seed medium: glucose 30.0 g/L, protein 10.0 g/L, enzyme powder 5.0 g/L, KH2PO4 4.5 g/L, CaCO3 2.0 g/L, pH 6.7.
the basic medium of pear slag fermentation: fresh pear slag 50 g, bran 20 g.
1.2 Main test instrumentZHY-2102C thermostatic culture oscillator (Shanghai Zhicheng Analytical Instrument Manufacturing Co., Ltd.), SPX-250B-II biochemical incubator (Shanghai Hedeshi Laboratory Equipment Co., Ltd.), HW. SY-P1 type electric heated water bath pot (Beijing Oriental Jingrui Technology Development Co., Ltd.), SW-CJ-2FD type ultra-clean work station (Suzhou Antai Air Technology Co., Ltd.).
1.3 Preparation of 1.3.1 seed fluid, the 2 to 3 ring LB bevel preservation of polyviable spores D1 inoculated into a 250 mL triangular bottle containing a 100 mL seed medium, 30 degrees C, 200 r/min shaker oscillation culture 14 h.
1.3.2 Solid fermentation of polyviable Bacillus D1 mixed fresh pear slag with bran in different quality ratios, and the proportion of pear slag and bran was controlled to 10:1, 10:2, 10:4, 10:6, respectively, with water content of 70.47%, 65.70%, 58.20%, 52.57%, and the pear residue and bran (38%) were added as a control ratio.
are called 70 g above the above-mentioned cultivation base into a 500 mL beaker, wrapped in 8 layers of gauze, 115 degrees C autoclave 15 min.
to be cooled to room temperature (25 degrees C), access to 0.1 mL/g multi-stick Bacillus spore seed liquid, placed in a biochemical culture tank 30 oC constant temperature static culture 72 h.
during fermentation, stir the turn every 12 h with a sterile glass bar.
3 parallels per group.
timed sampling to monitor the growth of bacteria and observe the formation of spores.
1.3.3 The influence of culture time according to 10% inoculation volume, the multi-stick Spore seed liquid is connected to the basic medium of pear slag fermentation, the temperature of 30 degrees C is static culture, and the formation of spores is observed.
1.3.4 Plus the effect spores of nitrogen sources, urea 0.7%, sodium nitrate 0.5%, ammonium nitrate 0.5%, ammonium chloride 0.5%, soybean meal 7.0%, dephenol cotton seed protein powder 7.0% to the basic medium of spore fermentation of pear slag were added to the basic medium of pear slag fermentation, respectively, by adding urea 0.7%, sodium nitrate 0.5%, ammonium nitrate 0.5%, ammonium chloride 0.5%, soybean meal 7.0%, dephenol cotton seed protein powder 7.0%, to not add any nitrogen source as a control.
10% seed solution, the temperature of 30 degrees C constant lyation culture 120 h.
1.3.5 The effects of adding inorganic salts adjusted the basic medium of pear slag fermentation to 50 g of pear slag, 20 g of bran, and 4.9 g of soy meal powder.
added sodium chlorate 0.5%, calcium carbonate 0.5%, magnesium sulfate 0.5%, potassium dihydrophosphate 0.5%, manganese sulfate 0.3%, iron sulfate 0.1%, respectively, to the above-mentioned basic medium of pear slag fermentation, respectively, to not add any inorganic salts as a control.
10% seed solution is connected, the temperature of 30 degrees C constant lyation culture 120 h.
1.3.6 The influence of culture temperature The composition of the pear slag fermentation medium was adjusted to the pear slag 50 g, bran 20 g, soybean meal powder 4.9 g, potassium dihydrophosphate 0.37 g.
connected to 10% seed fluid, the temperature and static culture at 25, 30, 34, 37 and 40 degrees C are heated at 120 h.
1.3.7 The impact of bottling is 50 g of pear slag, 20 g of bran, 4.9 g of soybean meal, 0.37 g of potassium dihydrophosphate.
are called 30, 50, 70, 90 and 110 g of the above-mentioned pear slag fermentation medium into a 500 mL beaker, the thickness of the culture layer is 1.5, 3.0, 4.7, 5.3 and 6.4 cm, respectively, wrapped in 8 layers of gauze, sterilization.
access to 10% multi-stick Bacillus spore seed fluid, at a constant temperature of 30 degrees C static culture 120 h.
1.3.8 Multi-viscosus spore D1 Most suitable fermentation conditions for verification of pear slag fermentation medium composition is pear slag 50 g, bran 20 g, soybean meal powder 4.9 g, potassium dihydrophosphate 0.37 g.
said to take 70 g of the above-mentioned pear slag fermentation culture base into a 500 mL beaker, the thickness of the culture substrate layer 4.7 cm, wrapped in 8 layers of gauze, sterilized.
access to 10% multi-stick Bacillus spore seed fluid, 34 degrees C constant temperature static culture 120 h.
1.3.9 Counting method of active bacteria and spores: using flat colony counting method to determine the number of live bacteria in the sample (cfu/g, in wet mass).
spore counting method: the to-test counting sample 80 oC water bath heating 15 min, and then using the flat colony counting method to count, the result of the count is the number of spores (cfu/g, in wet mass). the
spore rate is the percentage of spores as a percentage of the number of active bacteria.
1.4 Data Processing Methods 3 parallels per group, variables are expressed in average-standard deviation (x-s), and each set of results is F-tested using Sps19.0 software to verify the significance of the differences between each group.
2 Results and analysis of 2.1 solid fermentation of polyviable Bacillus D1 This paper uses screw-type juicer to produce fresh duck pear slag is loose bean curd slag, suitable for use as a solid fermentation medium.
but fresh pear slag has a high water content of 76.2%, which may not be conducive to solid fermentation of polyviable Bacillus D1.
bran is a by-product of wheat flour processing, low water content, containing more starch, protein and other nutrients, and low price.
therefore, when the saiton of polyviable Bacillus Spores D1 is fermented, different proportions of bran are added to the fresh pear slag, the purpose of which is to regulate the moisture content of the pear slag on the one hand and the nutrition of the pear slag on the other, the results of which are shown in Figure 1.
note: fermentation temperature of 30 degrees C, fermentation time 72 h.
the difference between the different letters represents a significant level of difference between groups (P.lt;0.05), the same as below.
Note: fermentaion temperature 30 c, ferment time 72 h. letters means the subject of the lot difference and test groups at 0.05 level, The same below. Figure 1 Multi-sticky spore D1 solid fermentation Fig.1 Solid-state fermentation of P.polymyxa D1 as shown in Figure 1, when fresh pears Slag (without any auxiliary material) for fermentation media, multi-stick Spore D1 bacteria growth is good, its number of live bacteria reached 1.6 x 109 cfu/g, with pure bran to do the culture of the medium when the number of live bacteria is very close, which shows that the pear slag nutrition is more abundant, is a good culture multi-stick Spore bacteria culture.
can also be seen from Figure 1, with the increase in the amount of bran added (10:1, 10:2, 10:4, 10:6), the aquaculture moisture content of the medium also decreased (70.47%, 65.70%, 58.20%, 52.57%), the growth of bacteria has been significantly improved (P.lt;0.05), the number of live bacteria increased significantly.
may be the cause of this phenomenon, the addition of bran not only reduces the moisture content of the medium, increases the oxygen supply, but also improves the nutrition of the medium.
when the ratio of pear residue to bran is 10:4, the maximum number of live bacteria reaches 2.5 x 109 cfu/g.
when the amount of bran added continues to increase, the number of live bacteria does not increase, so the appropriate quality ratio of pear slag to bran is 10:4.
during 72 h fermentation, the formation of spores was examined and no spores formed.
because the formation of spores is very conducive to the production, storage and use of microbial agent, spore rate is an important indicator of microbial agent quality, therefore, how to make a large number of spores produced by the large number of spores of the many viscous spores become the focus of follow-up research.
2.2 The effect of fermentation conditions on polystick spores D1 production spores 2.2.1 culture time is one of the important factors affecting the formation of spores of Bacillus spores.
as can be seen from Figure 2, the number of live bacteria as the incubation time increases.