Peach gum polysaccharide capable of efficiently resisting allergy and relieving itching, fermentation extraction method and application thereof
A technology of peach gum polysaccharide and extraction method, which is applied in the field of bioengineering, can solve the problems of poor uniformity and wide molecular weight distribution of products, and achieve the effects of low emission, narrow molecular weight range, and simple formula operation
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Embodiment 1
[0033] Embodiment 1 Preparation of engineering yeast dry powder
[0034] 1) Preparation of yeast clones. According to the amino acid sequence of 1,3-galactanase (sequence from NCBI: GenBank: KAA8623185.1), a DNA sequence was synthesized, digested and ligated with nucleases, and obtained linearity without signal peptide. The linearized DNA, DNA synthesis and vector construction were purchased from Shanghai Nupu Biology, and the linearized DNA was dissolved in 5-10 μL TE. Take 80 μL of commercially competent Pichia pastoris GS115 (purchased from Tiangen Biology) and mix it with 10 μg of linearized DNA, and transfer it into a pre-cooled 0.2 cm electroporation cup. Place on ice for 5 min. Set the machine parameters, immediately add 1ml of pre-cooled 1M sorbitol to the cup, transfer the contents to a sterilized centrifuge tube, divide it into 200μL aliquots, spread it on the MD plate (basic glucose medium), and incubate the plate at 30°C to cloning.
[0035] 2) Expression and pu...
Embodiment 2
[0036] Example 2 Extraction of Peach Gum Polysaccharide by High Pressure Water Extraction
[0037] After the peach gum is sorted out of debris, take about 100g of peach gum and add appropriate amount of water to soak overnight, add ultra-pure water according to the ratio of material to liquid 1:20, stir and extract with hot water at 80°C, extraction time is 4 hours, and the number of times is 2 times. Due to the high viscosity of the solution, first filter it with 2 layers of gauze, then centrifuge the filtrate at 8000r / min for 15 minutes, then pour out the supernatant, reduce the pressure and rotary evaporate to about 1 / 3 of the original volume, pour it out, and add absolute ethanol to The mass fraction of ethanol was 90%, and the supernatant was poured off after standing overnight. The lower precipitate was washed with absolute ethanol and vacuum filtered, then freeze-dried to obtain peach gum polysaccharide. The total sugar content of peach gum polysaccharides was determin...
Embodiment 3
[0041] Example 3 Extraction of Fermented Peach Gum Polysaccharide
[0042] After the peach gum is removed from debris, take about 100g of peach gum and add appropriate amount of water to soak overnight, and add ultra-pure water according to the material-liquid ratio of peach gum, ultrapure water and yeast dry powder expressing galactanase 1:10:0.01 and dry yeast powder (galactanase expressed), then poured into a 5L fermenter, added 0.5% methanol, adjusted the temperature to 30°C, rotated at 250rpm, stirred and fermented for 10h. Measure the viscosity of the feed liquid, and stop the fermentation when the viscosity drops below 500mPa.s. Pour out feed liquid, centrifuge 10 minutes with high-speed centrifuge 6000rpm. Discard the bacterial sediment at the bottom, and perform microfiltration using a roll filter membrane (pore size 0.22 microns) to remove residual bacterial cells and particulate components without causing bacterial contamination. Add absolute ethanol until the mas...
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