CN109749955B

CN109749955B - Lactobacillus brevis ZJ401 and application thereof

Info

Publication number: CN109749955B
Application number: CN201811630743.1A
Authority: CN
Inventors: 吴石金; 邱乐泉; 张嘉琳; 钟卫鸿; 李彤彤
Original assignee: Zhejiang University of Technology ZJUT
Current assignee: Wu Chenghu
Priority date: 2018-12-29
Filing date: 2018-12-29
Publication date: 2020-08-21
Anticipated expiration: 2038-12-29
Also published as: CN109749955A

Abstract

The invention provides a Lactobacillus brevis ZJ401 with antioxidant activity and application thereof, wherein the strain is preserved in China center for type culture Collection with the address as follows: china, wuhan university, zip code 430072, deposit number: CCTCC M2017666, date of deposit: 11 and 7 months in 2017. The strain can obviously improve SOD and CAT activities in A549 cells, obviously reduce LDH in cell supernatant and ROS content in cells, and obviously improve the expression of Nrf2 and gamma-GCS genes in Caco-2 cells. The Lactobacillus brevis ZJ401 can be made into antioxidant food or health product for human body after being made into powder, and can be directly eaten or drunk with water, with convenient use and immunity enhancing effect. The lactobacillus brevis ZJ401 has important research significance and application prospect for the development of microecological viable bacteria preparations and functional fermented dairy products.

Description

Lactobacillus brevis ZJ401 and application thereof

(I) technical field

The invention relates to a strain-Lactobacillus brevis ZJ401 with in vivo and in vitro cell anti-oxidative stress injury activity and application thereof.

(II) background of the invention

The uk scholars, hamna, 1956, first proposed the theory of free radical aging, which theory of free radical aging (FRTA) postulated that aging changes are caused by free radical reactions. The 1990 united states aging research authority professor Sohal pointed out the deficiency of the theory of free radical aging and proposed for the first time this concept of oxidative stress. Oxidative stress, i.e., increased production or decreased scavenging ability of free radicals in the body, causes disorders in the oxidative and antioxidant systems of the body, resulting in oxidative damage processes caused by accumulation of free radicals in the body. Not only is oxidative stress closely related to aging, but research has also shown that oxidative stress is closely related to the development of tumors, asthma, depression, and various chronic inflammatory diseases. Oxidative stress is also involved in the occurrence and development of cardiovascular diseases, which lead to atherosclerosis, heart failure, hypertension, myocardial damage and other cardiovascular diseases.

The body has limited antioxidant capacity, and therefore, the intake of exogenous antioxidants becomes an important way for the body to relieve oxidative stress. Lactic acid bacteria are a probiotic widely used by humans, often in the food fermentation industry and in micro-ecological preparations. As is well known, lactic acid bacteria have the functions of regulating intestinal flora, improving human immunity and the like. Some lactic acid bacteria have been found to be capable of scavenging active oxygen free radicals such as DPPH and hydroxyl free radical, and relieving oxidative stress. Therefore, the functional lactic acid bacteria can be used as a natural food additive to relieve oxidative stress and the occurrence of some diseases. The screening of the lactobacillus with the function of relieving the oxidative stress has important significance and market value. According to the invention, the lactobacillus capable of relieving oxidative stress is screened from the traditional self-made fermented food through an in vitro antioxidant experiment, so that a foundation and a theoretical basis are laid for developing the lactobacillus food leavening agent with the function of relieving oxidative stress.

Disclosure of the invention

The invention aims to provide a novel strain, namely Lactobacillus brevis ZJ401, which has the activity of resisting oxidative stress damage of cells in vivo and in vitro.

The technical scheme adopted by the invention for achieving the aim of the invention is as follows:

the invention provides a new strain-Lactobacillus brevis ZJ401, which is preserved in China center for type culture Collection with the preservation date: 11/7/2017, accession number: CCTCC No: m2017666, address: china, wuhan university, zip code 430072. The invention obtains a pure culture with antioxidant function from pickled vegetables through primary screening and secondary screening, and the strain is determined to be Lactobacillus brevis (Lactobacillus brevis) named Lactobacillus brevis ZJ401(Lactobacillus brevis ZJ401) through morphological identification, physiological and biochemical test identification and 16S rDNA sequence analysis.

The invention also relates to application of the lactobacillus brevis ZJ401 in preparation of a cell antioxidant, wherein the cell is human lung cancer cell A549 or human colon cancer cell Caco-2, and the cell antioxidant can improve SOD and CAT activities in the cell, reduce LDH in cell supernatant and ROS content in the cell, and increase expression levels of Nrf2, SOD and HO-1 genes in the cell.

The cell antioxidant is wet thallus obtained by fermenting and culturing lactobacillus brevis ZJ401 or supernatant obtained by ultrasonically crushing the wet thallus, and is prepared by the following method:

(1) inoculating lactobacillus brevis ZJ401 to an MRS solid culture medium, and culturing at 37 ℃ for 24h to obtain slant thallus; MRS solid medium: 10.0g/L of peptone, 10.0g/L of beef extract, 10.0g/L of yeast extract, 2.0g/L of diammonium citrate, 5.0g/L of sodium acetate, 0.58g/L of magnesium sulfate, 0.25g/L of manganese sulfate, 2.0g/L of diammonium hydrogen phosphate, 20.0g/L of glucose, 801mL/L of tween-801, 18g/L of agar, deionized water as a solvent and 5.0 of pH value;

(2) inoculating the slant thallus to an MRS liquid culture medium, and culturing for 16h at 37 ℃ to obtain a seed solution; the MRS liquid culture medium is agar in the MRS solid culture medium;

(3) inoculating the seed liquid into MRS liquid culture medium with the inoculation amount of 3% of volume concentration, and performing fermentation culture at 37 ℃ to obtain fermentation liquid containing wet thalli, namely complete cell suspension with the concentration of 10⁹cfu/kg; and (3) ultrasonically crushing 5ml of the complete cell suspension, wherein the power is 400W, the work is 5s, the stop is 5s, the time is 60 times, centrifuging the bacterial suspension for 10min at the temperature of 4 ℃ and the speed of 10000r/min after the ultrasonic crushing, and collecting 4ml of supernatant to obtain the cell-free extract.

The invention also provides an application of the lactobacillus brevis ZJ401 in preparing an antioxidant feed, wherein the content of the lactobacillus brevis ZJ401 in the antioxidant feed is 10⁹cfu/kg or more, preferably 10⁹cfu/kg。

The invention also provides a method for preparing the lactobacillus brevis ZJ401Application of lactobacillus brevis ZJ401 in antioxidant food or health care product with content of 10⁹cfu/kg or more.

The invention has the following beneficial effects: the lactobacillus brevis ZJ401 with antioxidant activity is obtained by screening, and has high antioxidant activity and good acid and bile salt resistance. The DPPH clearance of the cell-free extract of ZJ401 was 25.68%, the hydroxyl radical clearance was 23.72%, and the reducing activity was 104.67 μmol/L L-cysteine. The fermentation liquor and the cell-free extract of ZJ401 can obviously improve SOD and CAT activities in A549 cells and obviously reduce LDH in cell supernatant and ROS content in the cells. In a Caco-2 oxidative stress model, the total antioxidant capacity of culture supernatant can be remarkably improved, and the expression of Nrf2 and gamma-GCS genes in Caco-2 cells can be remarkably improved by the complete cell suspension of ZJ 401. The Lactobacillus brevis ZJ401 can be made into antioxidant food or health product for human body after being made into powder, and can be directly eaten or drunk with water, with convenient use and immunity enhancing effect. The lactobacillus brevis ZJ401 has important research significance and application prospect for the development of microecological viable bacteria preparations and functional fermented dairy products.

(IV) description of the drawings

FIG. 1 is colony morphology of strain ZJ401 on MRS solid medium;

FIG. 2 is a gram-stained microscopic picture of strain ZJ401 (red rod-shaped strain is negative control, purple rod-shaped strain ZJ 401);

FIG. 3 is a growth curve of strain ZJ 401;

FIG. 4 is a clade of strain ZJ 401;

FIG. 5 is the effect of strain ZJ401 on ROS production; a-blank control group; b-oxidative stress control group; C-FS test group; D-CFE experimental group; E-VC positive control group; .

FIG. 6 shows the effect of strain ZJ401 on the expression of mRNA of oxidative stress-related genes Nrf2(a), SOD (b), HO-1(c), and gamma-GCS (d) in Caco-2 cells; CFE: a cell-free extract; IC: a suspension of intact cells; a to e represent significance analysis among different groups in the same gene, the same letters represent no significance, and the different letters represent significance (p < 0.05).

(V) detailed description of the preferred embodiments

The invention will be further described with reference to specific examples, but the scope of the invention is not limited thereto:

example 1: screening and identification of Lactobacillus brevis ZJ401

1. Screening of Strain ZJ401

(1) Primary screening of strains: collecting pickle soup prepared by a traditional fermentation method by using a disposable sterile syringe, carrying out gradient dilution on a sterile collected sample by using normal saline, uniformly coating 100-200 mu L of the diluted sample on an MRS solid culture medium with pH of 5.0, positively arranging the diluted sample in a 37 ℃ biochemical incubator for 30min, and inversely culturing the dried bacterial liquid for 24-48 h. And (3) observing the colony morphology of the single colony on the flat plate, and selecting the colony conforming to the lactic acid bacteria morphology (the colony is circular, medium-sized, convex, whitish, moist and neat in edge). Gram staining and catalase test were performed on the colonies after separation and purification. The strains with gram-positive staining and negative catalase test can be primarily judged as lactobacillus.

(2) And (3) tolerance screening: inoculating the activated strains twice with 3 vol% MRS liquid culture medium with pH of 2.0, 3.0, 4.0, 5.0, 6.0, 0, 0.3 wt% and 0.5 wt% of pig bile salt, H-containing MRS liquid culture medium₂O₂MRS liquid culture medium with concentration of 0.4, 0.7, 1.0mmol/L respectively. Culturing at 37 deg.C for 16h, and selecting the strain with the best tolerance by measuring absorbance at 600 nm. Screening to obtain the protein with the pH value of 3.0, the mass fraction of the pig bile salt of 0.5 percent and the content of H₂O₂The strain ZJ401 which keeps activity in the environment with the concentration of 1.0 mmol/L.

(3) Sample treatment, inoculating the strain ZJ401 in MRS liquid culture medium, culturing at 37 deg.C for 16h, centrifuging the bacterial liquid at 4 deg.C and 4000r/min for 10min, discarding the supernatant, collecting the thallus, washing the thallus with PBS buffer solution with pH of 7.2 for 3 times, resuspending, adjusting the number of the thallus to 1 × 10⁹cfu/mL is complete cell suspension; and (3) carrying out ultrasonic disruption on the whole cell suspension, wherein the power is 400W, the work is carried out for 5s, and the stop is carried out for 5s and 60 times. After ultrasonic crushing, the bacterial suspension is treated at the temperature of 4 ℃,centrifuging at 10000r/min for 10min, and collecting supernatant as cell-free extract.

(4) And (3) detecting and screening antioxidant activity: and (3) detecting DPPH clearance rate, hydroxyl radical clearance rate, lipid peroxidation resistance rate, ferrous ion chelating capacity and reduction activity of the whole cell suspension and the cell-free extract sample, and detecting SOD, GSH-Px and NADH oxidase activity of the cell-free extract. DPPH clearance rates of the extracts of the intact cells and the acellular extracts of ZJ401 are respectively 19.33 percent and 25.68 percent, hydroxyl radical clearance rates are respectively 19.82 percent and 23.72 percent, anti-lipid peroxidation capacities are respectively 12.11 percent and 8.62 percent, ferrous ion chelating capacities are respectively 12.61 percent and 15.43 percent, and reduction activities are respectively equivalent to the amounts of 97.00 mu mol/L and 104.67 mu mol/L L-cysteine. The SOD activity of ZJ401 is 7.068U/mgprot, the GSH-Px activity is 156.73U/mgprot, and the NADH oxidase activity is 0.0615U/mgprot.

MRS liquid medium: 10.0g/L of peptone, 10.0g/L of beef extract, 10.0g/L of yeast extract, 2.0g/L of diammonium citrate, 5.0g/L of sodium acetate, 0.58g/L of magnesium sulfate, 0.25g/L of manganese sulfate, 2.0g/L of diammonium hydrogen phosphate, 20.0g/L of glucose and 801mL/L of tween, wherein a solvent is deionized water, and the pH value is 5.0.

The MRS solid culture medium is prepared by adding 18g/L agar into an MRS liquid culture medium.

2. Identification of Strain ZJ401

(1) And (3) colony morphology identification: the strain ZJ401 is inoculated on an MRS solid medium and cultured for 24h at 37 ℃, and a single colony is in a whitish, convex and smooth and moist circle shape (shown in figure 1). Under microscopic examination, the strain ZJ401 is gram-positive, and the shape of the strain ZJ401 is straight or slightly bent and short rod-shaped, and sometimes single, sometimes paired or chain-shaped (as shown in figure 2). The strain ZJ401 grows in a logarithmic phase at 6h, the thallus grows in a stationary phase after 18h, the logarithmic phase of the strain ZJ401 is 6-18 h, and the pH of the fermentation liquor is finally reduced to 4.4 from the initial 6.0 (as shown in figure 3).

(2) Physiological and biochemical identification: the strain ZJ401 is inoculated on MRS solid culture medium, cultured for 24h at 37 ℃, transferred into API50CHL culture medium, and prepared into bacterial suspension with 2 Mai's concentration by utilizing a Mai's turbidimeter tube. Inoculating the bacterial suspension into an API50CH reagent strip, and carrying out anaerobic culture at 37 ℃ for 24-48 h. The physiological and biochemical results show that the strain ZJ401 is Lactobacillus brevis at 24h and 48h (the specific physiological and biochemical results are shown in Table 1).

TABLE 1 physiological and biochemical results of Strain ZJ401

(3) And (3) molecular identification: the DNA of strain ZJ401 was amplified by PCR to obtain a 1455bp DNA fragment (SEQ ID NO. 1) using bacterial 16S rDNA universal primers. Sequences were BLAST aligned at NCBI. The alignment result showed that the strain with the highest sequence homology of the strain ZJ401 was Lactobacillus brevis (similarity 99%, SEQ ID NO: NR 116238.1). Selecting typical strain sequences with high homology from NCBI, and calculating genetic distance between the strain ZJ401 and the strain with similar development relation. Phylogenetic trees were constructed using MAGE 5.0 software (as shown in figure 4). Combining the form, the 16S rDNA sequence comparison result and the physiological and biochemical measurement result, determining that the strain ZJ401 is Lactobacillus brevis (Lactobacillus brevis), is named as Lactobacillus brevis ZJ401(Lactobacillus brevis ZJ401), is preserved in the China center for type culture Collection, and has the preservation number: CCTCC No: m2017666, date of deposit: 11/7/2017, address: china, wuhan university, zip code 430072.

Example 2 Lactobacillus brevis ZJ401 fermentation broth

(1) And inoculating lactobacillus brevis ZJ401 to an MRS solid culture medium, and culturing at 37 ℃ for 24h to obtain slant thallus.

(2) Inoculating the slant thallus to MRS liquid culture medium, and culturing at 37 deg.C for 16h to obtain seed liquid.

(3) Inoculating the seed liquid into MRS liquid culture medium with the inoculation amount of 3% of volume concentration, and performing fermentation culture at 37 ℃ to obtain fermentation liquid, namely complete cell suspension with the concentration of 10⁹cfu/kg; and (3) ultrasonically crushing 5ml of the complete cell suspension, wherein the power is 400W, the work is 5s, the stop is 5s, the time is 60 times, centrifuging the bacterial suspension for 10min at the temperature of 4 ℃ and the speed of 10000r/min after the ultrasonic crushing, and collecting 4ml of supernatant to obtain the cell-free extract.

Example 3 protective Effect of Lactobacillus brevis ZJ401 on oxidative stress injury of A549 cells in vitro

Preparation of PM2.5 solution: a sampling filter (Whatman PM2.5, from olgfsen) was cut to a size of 1cm × 1cm, immersed in ultrapure water, allowed to stand at room temperature for 30min, sonicated for 30min, and repeated 3 times to elute PM2.5 particles. The obtained filtrate was filtered with 6 layers of gauze, and the filtered filtrate was frozen at-80 ℃ to obtain a lyophilized powder. When in use, sterile PBS is added to prepare high-concentration mother liquor, and then the mother liquor is diluted into PM2.5 solutions with different concentrations.

PM2.5 solution was used as an oxidative stressor for A549 cells were cultured at 1 × 10 per well⁴Individual cell (i.e. concentration 1 × 10)⁵100. mu.L per well) were plated in 96-well plates containing 1640 basic medium containing 10% by volume calf serum + 1% by volume double antibody (100U/mL) and cultured at 37 ℃ for 24 hours, the medium was aspirated after the cells were attached to the walls, and after washing with PBS, the cells were grouped according to Table 2, PM2.5 was added to a final concentration of 100. mu.g/mL, the amount of the complete cell suspension (IC) of Lactobacillus brevis ZJ401 prepared in example 2 was 0.5mL, the amount of the cell-free extract (CFE) was 0.5mL, Vc was added to a final concentration of 20. mu.g/mL, and the same amount of PBS was used as a blank control and cultured at 37 ℃ for 24 hours. The fermentation liquor and the cell-free extract of the ZJ401 remarkably improve the SOD and CAT activities in the cells, remarkably reduce the contents of LDH in cell supernatant and ROS in the cells, and the ZJ401 has a relieving effect on oxidative stress damage of A549 cells caused by PM2.5 (as shown in Table 3 and figure 5).

TABLE 2 Experimental groups

TABLE 3 Effect of Lactobacillus brevis ZJ401 on SOD, CAT, LDH of A549 cells

Note: 1. in the table, a-d represent significance analysis among different groups under the same antioxidant enzyme index, the same letters represent no significance, and the different letters represent significance (p is less than 0.05).

CFE: a cell-free extract; IC: whole cell suspension

Example 4 protective Effect of Lactobacillus brevis ZJ401 on Caco-2 cell oxidative stress injury in vitro

By means of H₂O₂A549 cells as oxidative stressors for Caco-2 cells at 1 × 10 per well⁴Individual cell (i.e. concentration 1 × 10)⁵100 μ L per well) were plated in a 96-well plate containing DMEM basal medium with 20% fetal bovine serum by volume and 1% double antibody by volume (100U/mL) by blotting at 37 ℃ for 24 hours, the cells were aspirated after adherence, washed with PBS, and then grouped according to Table 4, where H is₂O₂The final concentration was 100. mu. mol/L, and the whole cell suspension (IC) of Lactobacillus brevis ZJ401 prepared in example 2 was added in an amount of 0.5ml and the cell-free extract (CFE) was added in an amount of 0.5 ml. The whole cell suspension and cell-free extract of ZJ401 significantly increased the total antioxidant capacity (T-AOC) in the cells (as shown in table 5).

TABLE 4 Experimental groups

TABLE 5 Effect of Lactobacillus brevis ZJ401 on Caco-2 antioxidant Activity

CFE: a cell-free extract; IC: whole cell suspension

Example 5 Effect of Lactobacillus brevis ZJ401 on intestinal epithelial Caco-2 cell Nrf2, SOD, HO-1, gamma-GCS mRNA expression in vitro

Caco-2 cells were purchased from ATCC. Removing the frozen cells fromTaking out the mixture from a refrigerator or a liquid nitrogen tank at minus 80 ℃, quickly placing the mixture into a water bath kettle at 37 ℃, shaking the mixture from time to melt the cells, adding 5mL of complete cell culture medium and unfrozen cell suspension into a centrifuge tube, centrifuging the mixture at 1000r/min for 3min, removing supernatant, adding complete culture medium to suspend the cells, fully blowing the cells by using a pipette, transferring the cell suspension into a cell culture dish, placing the cell culture dish into an incubator at 37 ℃ and 5% of carbon dioxide, replacing the culture solution once in 2 days, culturing the intestinal epithelium Caco-2 cells in vitro, and utilizing different dosages (2 × 10)⁵，2×10⁶，2×10⁷CFU) of the lactobacillus brevis ZJ401 respectively stimulates the small intestine epithelial cells for 8h, extracts the total RNA of the stimulated cells, carries out reverse transcription to obtain cDNA, carries out FQ-PCR (fluorescence quantitative polymerase chain reaction) to detect the mRNA expression changes of the intestinal epithelial cells Nrf2, SOD, HO-1 and gamma-GCS (gamma-GCS), and takes PBS (phosphate buffer solution) as a blank control, as shown in figure 6. The results show that the oxidative stress group has significantly increased expression levels (p) of Nrf2, SOD and HO-1 genes compared with the blank control group<0.05), described in H₂O₂The stimulation of the protein can lead Caco-2 cells to generate oxidative stress, and a Keap1-Nrf2-ARE signal channel in the cells is activated, and antioxidant enzyme genes such as SOD, HO-1 and the like in the downstream ARE activated.

Example 6 Effect of addition of Lactobacillus brevis ZJ401 to the diet on antioxidant function of mice

The Lactobacillus brevis ZJ401 can be added into the daily ration of 48-day-old mice to make the viable count reach 10⁹cfu/kg, continuously feeding for 21d, and detecting serum T-SOD and MDA. ZJ401 can significantly improve the activity (p-SOD) of 21d serum and intestinal mucosa total superoxide dismutase (T-SOD) of mice<0.05), reducing the content (p) of Malondialdehyde (MDA) in the 21d serum of the white mouse<0.01) and has a tendency of reducing the MDA content of 21d intestinal mucosa (p is 0.057), and can remarkably improve the T-SOD activity and the total antioxidant capacity (T-AOC) (p) of 42d serum<0.01)。

Sequence listing

<110> Zhejiang industrial university

<120> Lactobacillus brevis ZJ401 and application thereof

<160>1

<170>SIPOSequenceListing 1.0

<210>1

<211>1455

<212>DNA

<213> Lactobacillus brevis (Lactobacillus breris)

<400>1

cgtggcgatg ctatactgca gtcgaacgag cttccgttga atgacgtgct tgcactgatt 60

tcaacaatga agcgagtggc gaactggtga gtaacacgtg ggaaatctgc ccagaagcag 120

gggataacac ttggaaacag gtgctaatac cgtataacaa caaaatccgc atggattttg 180

tttgaaaggt ggcttcggct atcacttctg gatgatcccg cggcgtatta gttagttggt 240

gaggtaaagg cccaccaaga cgatgatacg tagccgacct gagagggtaa tcggccacat 300

tgggactgag acacggccca aactcctacg ggaggcagca gtagggaatc ttccacaatg 360

gacgaaagtc tgatggagca atgccgcgtg agtgaagaag ggtttcggct cgtaaaactc 420

tgttgttaaa gaagaacacc tttgagagta actgttcaag ggttgacggt atttaaccag 480

aaagccacgg ctaactacgt gccagcagcc gcggtaatac gtaggtggca agcgttgtcc 540

ggatttattg ggcgtaaagc gagcgcaggc ggttttttaa gtctgatgtg aaagccttcg 600

gcttaaccgg agaagtgcat cggaaactgg gagacttgag tgcagaagag gacagtggaa 660

ctccatgtgt agcggtggaa tgcgtagata tatggaagaa caccagtggc gaaggcggct 720

gtctagtctg taactgacgc tgaggctcga aagcatgggt agcgaacagg attagatacc 780

ctggtagtcc atgccgtaaa cgatgagtgc taagtgttgg agggtttccg cccttcagtg 840

ctgcagctaa cgcattaagc actccgcctg gggagtacga ccgcaaggtt gaaactcaaa 900

ggaattgacg ggggcccgca caagcggtgg agcatgtggt ttaattcgaa gctacgcgaa 960

gaaccttacc aggtcttgac atcttctgcc aatcttagag ataagacgtt cccttcgggg 1020

acagaatgac aggtggtgca tggttgtcgt cagctcgtgt cgtgagatgt tgggttaagt 1080

cccgcaacga gcgcaaccct tattatcagt tgccagcatt cagttgggca ctctggtgag 1140

actgccggtg acaaaccgga ggaaggtggg gatgacgtca aatcatcatg ccccttatga 1200

cctgggctac acacgtgcta caatggacgg tacaacgagt tgcgaagtcg tgaggctaag 1260

ctaatctctt aaagccgttc tcagttcgga ttgtaggctg caactcgcct acatgaagtt 1320

ggaatcgcta gtaatcgcgg atcagcatgc cgcggtgaat acgttcccgg gccttgtaca 1380

caccgcccgt cacaccatga gagtttgtaa cacccaaagc cggtgagata accttcggga 1440

gtcagccgtc taagt 1455

Claims

1. Lactobacillus brevis (Lactobacillus brevis) ZJ401, deposited in China center for type culture Collection with the deposit number: CCTCC No: m2017666, date of deposit: 11/7/2017, address: china, wuhan university, zip code 430072.

2. The use of Lactobacillus brevis ZJ401 as claimed in claim 1 for preparing a cellular antioxidant, wherein the cellular antioxidant is a fermentation broth obtained by fermentation culture of Lactobacillus brevis ZJ401 or a supernatant after ultrasonication of the fermentation broth.

3. Use according to claim 2, characterized in that the cell is a human lung cancer cell a549 or a human colon cancer cell Caco-2.

4. Use according to claim 2, characterized in that the cellular antioxidant is an agent that increases the activity of SOD or CAT in cells.

5. Use according to claim 2, characterized in that the cellular antioxidant is an agent that reduces the level of LDH or ROS in the cell.

6. The use according to claim 2, wherein the cellular antioxidant is an agent that increases the expression level of Nrf2, SOD or HO-1 genes in cells.

7. Use of Lactobacillus brevis ZJ401 as claimed in claim 1 for preparing an antioxidant feed.

8. The use according to claim 7, characterized in that the feed has a Lactobacillus brevis ZJ401 content of 10⁹cfu/kg or more.

9. Use of the Lactobacillus brevis ZJ401 as claimed in claim 1 in preparing an antioxidant food or health care product.

10. The use according to claim 9, characterized in that the antioxidant food or health product has a lactobacillus brevis ZJ401 content of 10⁹cfu/kg or more.