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Bacillus subtilis and method for producing nano-selenium by using bacillus subtilis
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A technology of Bacillus subtilis and nano selenium, which is applied in the field of microbial fermentation, can solve problems such as non-precipitation, achieve good antioxidant capacity and relieve cellular oxidative stress.
Active Publication Date: 2022-07-01
ZHEJIANG UNIV
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For example, low-speed centrifugation aims to remove bacteria by centrifugation to retain nano-selenium in the supernatant, but this method cannot completely precipitate proteins, organic selenium or polysaccharides, so the particle size of the supernatant product is relatively large, often reaching hundreds or even thousands of nanometers ;N-hexane extraction and centrifugation method aims to separate the substances dissolved in the organic phase in the product from the water phase by n-hexane extraction, and further collect the water phase and remove the bacteria by centrifugation, but it is also similar to the low-speed centrifugation method, and the supernatant of the water phase The particle size of the product is still hundreds of nanometers
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[0030] LB plate medium: NaCl 10g, tryptone 10g, yeast extract 5g, agar 25g, deionized water 1L, sterilized by high temperature and high pressure at 121°C for 21min.
[0031] Seed medium: 10 g of glucose, 5 g of tryptone, 5 g of yeast extract, 5 g of NaCl, 1 L of deionized water, sterilized by high temperature and high pressure at 115°C for 30 min.
[0032] Fermentation medium: 30g sucrose, 10g tryptone, 15g yeast extract, K 2 HPO 4 1g, NaCl 5g, MgSO 4 1.5g, 1L deionized water, pH 7.0, sterilized by high temperature and high pressure at 115°C for 30min.
[0033] 2. Strain activation
[0034] Bacillus subtilis frozen at -80°C B. subtilis The SR41 strain was thawed at room temperature, a loop was streaked on LB plate medium, and cultured at 37°C for 12h. Pick a single colony from the plate medium, inoculate it in the seed medium, and cultivate it at 37°C a...
Embodiment 2
[0040] Example 2 Characterization of Bacillus subtilis and nano-selenium complex
[0041] The Bacillus subtilis and nano-selenium complex in Example 1 was taken and fixed with 2.5% glutaraldehyde for 12 hours. Washed with PBS, fixed with 1% osmic acid, dehydrated with alcohol gradient, treated with isoamyl acetate, dried at critical point and coated, and observed under field emission scanning electron microscope.
[0042] The result is as figure 2 shown, Bacillus subtilis B. subtilis SR41 reduces sodium selenite to form nano-selenium, and releases it to the outside of the cell to form a complex of Bacillus subtilis and nano-selenium, in which nano-selenium is spherical, which is indicated by the arrow in the figure.
Embodiment 3
[0043] Example 3 Extraction method and particle size comparison of nano-selenium in the composite
[0044] Get the Bacillus subtilis and the nano-selenium complex in Example 1, respectively adopt modes such as ultrasonic crushing, low-speed centrifugation, n-hexane extraction and centrifugation, and aim to separate the Bacillus subtilis and nano-selenium to extract nano-selenium, and carry out particle size. detection.
[0045] Ultrasonic crushing method: take the fermentation broth in a centrifuge tube, place it in an ultrasonic crusher, sonicate for 5s on / 5s off, 60HZ, power 100W, sonicate in an ice bath for 20min, and use it for particle size detection;
[0046] Low-speed centrifugation method: take the fermentation broth in a centrifuge tube, place it in a centrifuge, 3000×g, centrifuge for 5 minutes, discard the supernatant, resuspend the precipitate in PBS, and use it for particle size detection;
[0047] N-hexane extraction and centrifugation method: take the fermentat...
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Abstract
The invention discloses bacillus subtilis and a method for producing nano-selenium by applying the bacillus subtilis. The bacillus subtilis SR41 is preserved on October 11, 2021, the preservation number of the bacillus subtilis SR41 is CGMCC (China General Microbiological Culture Collection Center) No.23536, and the bacillus subtilis SR41 has the capabilities of efficiently reducing sodium selenite and generating nano-selenium. After the bacillus subtilis SR41 is subjected to selenium-rich fermentation, a compound of the bacillus subtilis and the nano-selenium can be obtained, and the nano-selenium in the compound can be further extracted, namely, the bacillus subtilis produces the nano-selenium. The invention discloses a preparation method for extracting the nano-selenium produced by the bacillus subtilis only through ultrasonication for the first time, and the average particle size of the extracted nano-selenium is 65.07 + / -2.39 nm. Compared with the existing nano-selenium extraction methods such as a low-speed centrifugation method or a normal hexane extraction and centrifugation method, the particle size of the obtained nano-selenium is lower, and the operation is simpler. Meanwhile, the nano-selenium produced by the bacillus subtilis disclosed by the invention has a function of relieving oxidative stress of cells, so that the nano-selenium has relatively good oxidation resistance.
Description
technical field [0001] The invention relates to the technical field of microbial fermentation, in particular to a Bacillus subtilis and a method for producing nano-selenium by using the same. Background technique [0002] Selenium is an essential trace element for animals, which participates in various metabolic pathways in the body and is essential for health. Studies have shown that selenium has the functions of anti-cancer and anti-oxidation, improving immunity, and regulating metabolism. [0003] In recent years, nano-selenium has attracted widespread attention as a new type of selenium product with a particle size between 1 and 100 nm, which makes it highly biologically active. However, the industrial synthesis of nano-selenium is often prepared by chemical methods, that is, adding reducing agents such as vitamin C to selenate or selenite solution, and additionally adding stabilizers such as bovine serum albumin. This process not only consumes resources, but also easi...
Claims
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