CN117535371A

CN117535371A - Method for improving astaxanthin yield of rhodozyma by using fungus exciton

Info

Publication number: CN117535371A
Application number: CN202311529019.0A
Authority: CN
Inventors: 杜新永; 刘军伟; 罗成盛; 李欣; 裴中鲜
Original assignee: Qingdao Runda Biotechnology Co ltd
Current assignee: Qingdao Runda Biotechnology Co ltd
Priority date: 2023-11-16
Filing date: 2023-11-16
Publication date: 2024-02-09

Abstract

The invention provides a method for improving the astaxanthin yield of rhodozyma by using a fungus exciton, which comprises the following steps: s1, preparing Phaffia rhodozyma seed solution; s2, preparing a fungus exciton: respectively taking fungus raw materials, adding pure water, centrifuging for several times, mincing, collecting the crushed materials, freezing, thawing, freeze-drying, crushing the freeze-dried materials, sieving, and sterilizing under high pressure steam to obtain fungus excitons; s3, fermenting Phaffia rhodozyma: adding the fungus exciton obtained in the step S2 into the Phaffia rhodozyma fermentation medium, then inoculating the Phaffia rhodozyma seed liquid obtained in the step S1, and culturing in a shaking table to obtain a fermentation liquid; s4, extracting astaxanthin. According to the method for improving the astaxanthin yield of the rhodozyma by using the fungal exciton, provided by the invention, the astaxanthin content of the rhodozyma is improved by adding the fungal exciton into the rhodozyma fermentation medium, the astaxanthin accumulation in the rhodozyma thallus is increased, and the effect of improving the astaxanthin yield of the rhodozyma fermentation liquor is achieved.

Description

Method for improving astaxanthin yield of rhodozyma by using fungus exciton

Technical Field

The invention relates to the technical field of microorganisms, in particular to a method for improving the astaxanthin yield of Phaffia rhodozyma by using a fungus exciton.

Background

Astaxanthin is a ketocarotenoid, the chemical name of which is 3,3' -dihydroxy-4, 4' -diketo-beta, beta ' -carotene, red solid powder, which is fat-soluble, insoluble in water and soluble in organic solvents. Astaxanthin is a chain-breaking antioxidant, has extremely strong antioxidant capacity, can remove nitrogen dioxide, sulfide, disulfide and the like, can reduce lipid peroxidation, and effectively inhibits lipid peroxidation caused by free radicals. Meanwhile, the composition has various physiological effects of inhibiting tumor occurrence, enhancing immunity, removing free radicals in the body and the like, and has the functions of resisting cancer, preventing arteriosclerosis, coloring and the like.

Rhodotorula (xanthophyllomyces dendrorhous) is a basidiomycete yeast which is mainly single-cell and sometimes forms tandem structure and pseudohypha. When observed under a microscope, the shape of the fluorescent lamp is elliptical, and the fluorescent lamp is red or orange. Rhodotorula is a low-temperature type microorganism with low-temperature facultative cold, and the growth temperature is 4-28 ℃.

The existing production methods of astaxanthin comprise a chemical synthesis method, an artificial extraction method and a microbial fermentation method, but the chemical synthesis method has complicated steps, the control of reaction conditions is difficult, and harmful substances are generated in the synthesis process; the manual extraction method mainly adopts an acid-base alternation method, but the content of astaxanthin extracted at present is very low, the extraction of the astaxanthin with immature process causes difficulty, and the method is not suitable for large-scale industrialized production; in the microbial fermentation method, the main fermentation strain of astaxanthin is rhodotorula at present, and the microbial fermentation method is more and more paid attention to. However, the yield of astaxanthin in the fermentation product of Phaffia rhodozyma is not high, which seriously affects the yield of astaxanthin.

Therefore, the prior art needs to be solved.

Disclosure of Invention

The invention aims at overcoming the defects of the prior art and provides a method for improving the yield of phaffia rhodozyma astaxanthin by using a fungus exciton. Based on the problems, the invention increases the astaxanthin content of the rhodozyma fermentation medium by adding a fungus exciton into the rhodozyma fermentation medium, increases the astaxanthin accumulation amount in rhodozyma thallus, and achieves the effect of increasing the astaxanthin yield of the rhodozyma fermentation liquid.

In order to achieve the above object, the present invention is realized as follows:

a method for improving the astaxanthin yield of rhodozyma by using a fungal elicitor, comprising the following steps:

s1, preparing Phaffia rhodozyma seed liquid:

purifying phaffia rhodozyma freeze-dried powder in a PDA culture medium, selecting a single colony after purification, inoculating the single colony into test tube liquid culture, carrying out shake culture to obtain test tube seed liquid, inoculating the test tube seed liquid into shake flask culture liquid, and carrying out shake culture again to obtain phaffia rhodozyma seed liquid;

s2, preparing a fungus exciton:

respectively taking fungus raw materials, adding pure water, centrifuging for several times until the fungus raw materials are washed cleanly, mincing, collecting the smashed materials, then freezing, thawing, freeze-drying, smashing the freeze-dried materials, sieving, and sterilizing under high-pressure steam to obtain fungus excitons;

s3, fermenting Phaffia rhodozyma:

adding the fungus exciton obtained in the step S2 into a phaffia rhodozyma fermentation medium, then inoculating a certain amount of phaffia rhodozyma seed liquid obtained in the step S1, and culturing in a shaking table to obtain a fermentation liquid;

s4, extracting astaxanthin:

and (3) taking the fermentation broth obtained in the step (S3), centrifuging, collecting the precipitate, adding a solvent into the precipitate, carrying out ultrasonic oscillation, centrifuging again, and collecting the supernatant to obtain the astaxanthin extract.

The excitons may be derived from a variety of organisms such as plants, fungi, bacteria, etc., and may be classified into two classes, biological excitons and non-biological excitons, depending on the nature of the exciton. The biological exciton mainly refers to microorganism exciton, such as fungus spore, mycelium, fungus cell wall component, fungus culture filtrate, etc. Non-biological elicitors are signal factors that are not natural components of the cell but trigger the formation of phytochemicals. The fungus exciton is a specific chemical substance or signal from fungus, and can rapidly, specifically and selectively induce the expression of specific genes of the fungus in the interaction of the fungus and the fungus so as to activate specific secondary metabolic pathways and accumulate specific target secondary products. In the invention, in particular, marine rhodotorula and red yeast rice are used as fungi to prepare fungus excitons, which can effectively stimulate the fungi including rhodotorula to increase the amount of metabolites and accumulate the concentration and content of astaxanthin as secondary products.

The method for improving the astaxanthin yield of the rhodozyma by using the fungus exciton comprises the step of preparing a fungus material, wherein the fungus material is any one or two of saccharomycetes, mushroom fungus and red yeast rice.

The method for improving the astaxanthin yield of the rhodotorula by using the fungus exciton is characterized in that the microzyme is any one of rhodotorula oceanica and rhodotorula mucilaginosa. Preferably, the yeast is rhodotorula marinosa.

The method for improving the astaxanthin yield of the rhodotorula by using the fungus exciton is characterized in that the fungus is any one of coriolus versicolor, ganoderma lucidum and ganoderma atrum.

The method for improving the astaxanthin yield of the rhodotorula by using the fungus exciton comprises the step of using any one of rhodotorula, coriolus versicolor, red rice, a mixture of a marine rhodotorula fermentation liquid and a coriolus versicolor fermentation liquid, a mixture of coriolus versicolor and red rice and a mixture of marine rhodotorula thallus and red rice as a fungus raw material. Preferably, the fungus raw material is any one of rhodotorula ocellata, red yeast rice, a mixture of rhodotorula ocellata fermentation liquor and coriolus versicolor fermentation liquor, a mixture of coriolus versicolor and red yeast rice, and a mixture of rhodotorula ocellata thallus and red yeast rice. More preferably, the fungal raw material is a mixture of rhodotorula oceanica thallus and red yeast rice. Most preferably, the fungus raw material is marine rhodotorula thallus and red yeast rice with the mass ratio of 1:1.

The method for improving the astaxanthin yield of the rhodozyma by using the fungal elicitor comprises the following specific steps of: respectively taking fungus raw materials, adding pure water, centrifuging at 3000-5000 rpm/min for 10-20 min, centrifuging for several times until the fungus raw materials are washed cleanly, taking precipitates, stirring, collecting the crushed materials, freezing at-70 ℃ to-90 ℃ for 4-6 days, thawing in a water bath kettle at 50-70 ℃ for three times, repeatedly freezing and thawing, crushing the lyophilized materials, sieving with a 60-mesh sieve, and sterilizing for 20-50 min under 0.1Mpa high-pressure steam to obtain fungus elicitors. The method for preparing the fungal exciton is easy to implement and convert, and the fungal exciton prepared by the method improves the astaxanthin content in the rhodotorula during the fermentation process.

A method for increasing the astaxanthin yield of rhodozyma by using a fungal elicitor as described above, wherein the shake culture conditions in the step S1 are as follows: the temperature is 20-30 ℃, the rotating speed is 160-180 rpm/min, and the culture time is 3-5 days; the secondary shaking culture conditions in the step S1 are as follows: the temperature is 20-30 ℃, the rotating speed is 160-180 rpm/min, and the culture time is 2-4 days. Preferably, the shake culture conditions in the step S1 are as follows: the temperature is 25 ℃, the rotating speed is 170rpm/min, and the culture time is 4 days; the secondary shaking culture conditions in the step S1 are as follows: the temperature was 25℃and the rotation speed was 170rpm/min, and the cultivation time was 3 days.

Based on the same inventive concept, the invention also provides an astaxanthin preparation, which comprises the astaxanthin extracting solution prepared by the method for improving the yield of the phaffia rhodozyma astaxanthin by using the fungus exciton.

Based on the same inventive concept, the invention also provides a use of an astaxanthin formulation according to the above in feeding poultry.

Compared with the prior art, the invention has the following effects and advantages:

1. according to the invention, the fungal exciton is added into the rhodozyma fermentation medium to improve the astaxanthin content, so that the astaxanthin accumulation amount in the rhodozyma thallus is increased, and the effect of improving the astaxanthin yield of the rhodozyma fermentation liquid is achieved.

2. The application of the astaxanthin preparation in feeding poultry provided by the invention can effectively improve the oxidation resistance of poultry (such as broiler chickens).

Detailed Description

The following description of the embodiments of the present invention will clearly and fully describe the technical solutions of the embodiments of the present invention, and it is apparent that the described embodiments are only some embodiments of the present invention, not all embodiments. All other embodiments, which can be made by those skilled in the art based on the embodiments of the invention without making any inventive effort, are intended to be within the scope of the invention.

Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. The terminology used in the description of the invention herein is for the purpose of describing particular embodiments only and is not intended to be limiting of the invention. The term "and/or" as used in this specification includes any and all combinations of one or more of the associated listed items.

The experimental methods used in the following examples are conventional methods unless otherwise specified.

Materials, reagents and the like used in the examples described below are commercially available unless otherwise specified.

Rhodotorula is purchased from Beijing Bei Naviet Union biotechnology institute, numbered BNCC336673; rhodotorula marinic acid is purchased from Beijing north na-invasive joint biotechnology institute, numbered BNCC119738; red yeast rice was purchased from Shanghai Bai Si food development Co., ltd; coriolus versicolor is purchased from Minyuan bacterial industry; SOD detection kit is purchased from Nanjing built bioengineering institute; GSH detection kit is purchased from Nanjing's institute of biological engineering.

Example 1:

s1, preparing Phaffia rhodozyma seed liquid:

taking phaffia rhodozyma freeze-dried powder, purifying for 2 times in PDA culture medium, wherein the purifying culture temperature is 25 ℃ and the purifying culture time is 6 days. The single colony after purification is selected and inoculated in test tube liquid culture, shake culture is carried out, and the shake culture conditions are as follows: the temperature was 25℃and the rotational speed was 170rpm/min for 4 days. Inoculating the test tube seed liquid into shake flask culture liquid with 2% of inoculation amount, and shake culturing again under the following conditions: the temperature is 25 ℃, the rotating speed is 170rpm/min, the culture time is 3 days, and the rhodozyma seed liquid is obtained. The rhodozyma seed liquid culture solution comprises the following components: glucose 20g/L, yeast extract 5g/L, peptone 5g/L, potassium dihydrogen phosphate 2g/L and magnesium sulfate 2g/L.

S2, preparing a fungus exciton:

mixing marine rhodotorula and pure water according to the mass ratio of 1:50 respectively, centrifuging at 4000rpm/min for 15min, and centrifuging for several times until the supernatant is clear after washing and centrifuging. Collecting precipitate, respectively crushing with a universal crusher, and collecting crushed materials. Then placing the mixture at the temperature of minus 80 ℃ for 5 days, thawing the mixture, placing the mixture in a water bath kettle at the temperature of 60 ℃, and repeatedly freezing and thawing the mixture for three times. And freeze-drying on a vacuum freeze dryer, crushing the freeze-dried product, and sieving the crushed product with a 60-mesh sieve. Sterilizing under high pressure steam for 30min to obtain marine rhodotorula fungus exciton.

S3, culturing Phaffia rhodozyma:

adding the ocean rhodotorula fungus exciton into the rhodotorula fermentation culture medium, adding the addition amount of 0.02%, and then inoculating the rhodotorula seed liquid with the volume ratio of 2%. Culturing in a shaking table under the following conditions: the temperature is 25 ℃, the rotating speed is 170rpm/min, and the culture time is 4 days, so that the fermentation broth is obtained. The rhodozyma fermentation medium comprises the following components: glucose 20g/L, yeast extract 5g/L, peptone 5g/L, potassium dihydrogen phosphate 2g/L and magnesium sulfate 2g/L.

S4, extracting astaxanthin:

collecting more than 40mL of fermentation broth, centrifuging to collect precipitate, adding 3mL of dimethyl sulfoxide (DMSO) at 55deg.C, ultrasonically oscillating for 30min, adding 5mL of acetone, ultrasonically extracting for 20min, centrifuging at 6000rpm for 10min, and collecting supernatant to obtain astaxanthin extractive solution.

Example 2:

the procedure is as in example 1, except that the marine rhodotorula of example 1 is replaced with coriolus versicolor.

Example 3:

the procedure is as in example 1, except that the marine rhodotorula of example 1 is replaced by red yeast rice.

Example 4:

the operation was the same as in example 1 except that the rhodotorula marinus of example 1 was replaced with a fermentation broth of rhodotorula marinus and coriolus versicolor.

The preparation method of the fermentation liquor of the rhodotorula maritima and coriolus versicolor comprises the following steps: respectively obtaining a marine red yeast fermentation liquid and a coriolus versicolor mycelium fermentation liquid by liquid shake flask fermentation of marine red yeast and coriolus versicolor respectively, and then mixing the marine red yeast fermentation liquid and the coriolus versicolor fermentation liquid according to a mass ratio of 1:1, mixing to obtain the fermentation liquor of the marine rhodotorula and the coriolus versicolor.

Example 5:

the procedure is as in example 1 except that the marine rhodotorula of example 1 is replaced by a mixture of coriolus versicolor and red yeast rice, wherein the mass ratio of coriolus versicolor to red yeast rice is 1:1.

example 6:

the procedure is as in example 1 except that the rhodotorula marinus of example 1 is replaced by a mixture of rhodotorula marinus cells and red rice, wherein the mass ratio of rhodotorula marinus cells to red rice is 1:1.

comparative example 1:

the procedure is as in example 1, except that no exciton is added. The specific method comprises the following steps:

the extraction method of the phaffia rhodozyma astaxanthin comprises the following steps:

s11, preparing Phaffia rhodozyma seed liquid:

S12, culturing Phaffia rhodozyma:

inoculating Phaffia rhodozyma seed liquid with the volume ratio of 2%, and culturing in a shaking table under the following culture conditions: the temperature is 25 ℃, the rotating speed is 170rpm/min, and the culture time is 4 days, so that the fermentation broth is obtained. The rhodozyma fermentation medium comprises the following components: glucose 20g/L, yeast extract 5g/L, peptone 5g/L, potassium dihydrogen phosphate 2g/L and magnesium sulfate 2g/L.

S13, extracting astaxanthin:

collecting more than 40mL of fermentation broth, centrifuging to collect precipitate, adding 3mL of dimethyl sulfoxide (DMSO) at 55deg.C, ultrasonically oscillating for 30min, adding 5mL of acetone, ultrasonically extracting for 20min, centrifuging at 6000rpm for 10min, and collecting supernatant to obtain astaxanthin extract.

Test example 1: statistics of biomass

1. The experimental method comprises the following steps:

for detecting the biomass in the obtained fermentation broth, the fermentation broths of examples 1 to 6 and comparative example 1 were taken respectively after the completion of shake flask culture, and the biomass was measured, and the magnitudes of the relevant values were counted. The specific detection method comprises the following steps:

(1) Dilution operation: 8 sterile tubes each containing 4.5mL of water were prepared and mixed at a pressure of 10 ^-1 To 10 ^-8 Is numbered in the order of (2).

(2) 0.5mL of the cultured fungus night was aspirated with a pipetting tube and 10 was injected ^-1 In a tube of double dilution.

(3) And (3) blowing and sucking for three times by using a pipetting gun, so that bacterial liquid and water are fully and uniformly mixed.

(4) From 10 ^-1 0.5mL of the dilution was aspirated into a double-diluted tube, and 10 was injected ^-2 Multiple diluted test tubeIn the second step, the mixing operation is repeated.

(5) And so on until the dilution of the last tube is completed.

(6) In operation, the nozzle and pipette should be operated at a distance of 1-2 cm from the flame.

(7) 0.1mL of the solution was added dropwise to the surface of the agar medium.

(8) The bacterial liquid is evenly coated on the surface of the culture medium by a coater, and the culture dish can be rotated during coating, so that the bacterial liquid is evenly distributed.

(9) Culturing in a 26 ℃ incubator for 3 days after coating, and counting the size of the biomass.

2. Experimental results and analysis:

TABLE 1 biological measurement results

Different treatment groups	Biomass CFU/mL
		Fermentation broth of example 1	1.3×10 ⁷ CFU/mL
Fermentation broth of example 2	2.1×10 ⁷ CFU/mL
		Fermentation broth of example 3	2.3×10 ⁷ CFU/mL
Fermentation broth of example 4	1.6×10 ⁷ CFU/mL
		Fermentation broth of example 5	1.5×10 ⁷ CFU/mL
Fermentation broth of example 6	1.4×10 ⁷ CFU/mL
		Fermentation broth of comparative example 1	1.9×10 ⁷ CFU/mL

As can be seen from Table 1, the seven groups of fermentation broths have a biomass statistic of 10 ⁷ The order of CFU/mL is not very different, which indicates that the rhodotorula biomass in each group of fermentation broths is on the same order of magnitude, and the addition of the fungal elicitor has no effect on the rhodotorula biomass.

Test example 2: statistics of astaxanthin production

1. The experimental method comprises the following steps:

the astaxanthin extraction solutions provided in examples 1-6 and comparative example 1 have a volume ratio of 3:5 was blank, and absorbance values (i.e., OD 475) were measured at 475 nm.

2. Experimental results and analysis:

TABLE 2 astaxanthin production amount measurement results

Different treatment groups	Absorbance value size
		Astaxanthin extract according to example 1	0.14
Astaxanthin extraction liquid according to example 2	0.12
		Astaxanthin extraction liquid according to example 3	0.15
Astaxanthin extraction liquid according to example 4	0.15
		Astaxanthin extraction liquid according to example 5	0.18
Astaxanthin extraction liquid according to example 6	0.28
		Astaxanthin extraction solution according to comparative example 1	0.12

As can be seen from table 2, by measuring the astaxanthin absorbance values in the seven groups of red-french-wave fermentation broths, it was found that the mass ratio of the addition of rhodotorula gracilis to red yeast rice in the medium of example 6 was 1: when the mixture of 1 was used as a starter, the amount of astaxanthin produced by Phaffia rhodozyma was the highest as compared with the other groups, and the measured values of example 1 and examples 3 to 5 were higher than those of comparative example 1. Since the biomass of each treated rhodozyma mother liquor is similar as seen in table 1, this suggests that the increase in astaxanthin in the rhodozyma mother liquor is mainly due to its own astaxanthin accumulation, and further that the addition of fungal elicitors to the culture medium contributes to the accumulation of rhodozyma astaxanthin, with the addition of rhodozyma cells and red yeast rice mixtures as elicitors being optimal.

Test example 3: determination of the free radical scavenging Rate of hydroxyl groups

1. The experimental method comprises the following steps:

the astaxanthin extracts of examples 1-6 and comparative example 1 were subjected to hydroxyl radicalsThe clearance is measured by the following method: taking 1mL of astaxanthin extract, adding 800. Mu.L of ferrous sulfate (6 mmol/L) and 800. Mu.L of salicylic acid (6 mmol/L) into a test tube, shaking, and adding 0.1% H into the test tube ₂ O ₂ 50 mu L of the mixture is mixed evenly, the water bath is carried out at 37 ℃ for 30min, and the absorbance value of each reaction solution is measured at 510nm and is recorded as A1; 1mL of astaxanthin extract was taken in a test tube, 800. Mu.L of ferrous sulfate (6 mmol/L) and 800. Mu.L of salicylic acid (6 mmol/L) were added to the test tube, and H was added to the test tube after shaking ₂ Mixing O50 mu L uniformly, measuring absorbance value of each reaction solution at 510nm in 37 ℃ water bath for 30min, and marking as A2; taking distilled water as blank, taking 1mL distilled water into a test tube, adding 800 mu L ferrous sulfate (6 mmol/L) and 800 mu L salicylic acid (6 mmol/L) into the test tube, shaking, and adding 0.1% H into the test tube ₂ O ₂ Mixing 50 μl, measuring absorbance value of each reaction solution at 37deg.C water bath for 30min and 510nm, and recording as A0; and the clearance is calculated as follows. The test was repeated three times.

Clearance (%) = [ A0- (A1-A2) ]/a0×100.

2. Experimental results and analysis:

TABLE 3 determination of hydroxyl radical scavenger

As can be seen from table 3, the hydroxyl radical scavenging rate statistics of the astaxanthin extracts from different groups showed that the astaxanthin extract of example 6 had the highest hydroxyl radical scavenging rate, and that the astaxanthin extracts of examples 5, 3, 4 and 1 had hydroxyl radical scavenging rates higher than that of comparative example 1, indicating that the addition of fungal elicitors to the culture medium helped to increase the hydroxyl radical scavenging rate of astaxanthin in rhodotorula cells, wherein the mass ratio of rhodotorula cells to red yeast rice added was 1:1, the effect of the mixture is optimal.

Test example 4: efficacy verification of antioxidant Activity

1. The experimental method comprises the following steps:

three groups are used for verifying whether the addition of the fungus exciton in the culture of the rhodozyma is helpful for improving the oxidation resistance of the broiler chickens. The experimental animals are selected from freshly hatched broilers, fed for about 7 days, and are divided into the following three groups by adding red Phaffia yeast fermentation broth of different fungus excitons:

TABLE 4 antioxidant Activity efficacy different treatment groups

7 broiler chickens in each group are fed with corresponding Phaffia rhodozyma fermentation liquor or water according to the different groups, and the feeding addition amount is 0.5 mL/chicken per day. Feeding for five days, injecting 0.3mL of escherichia coli into each chicken to enable the chickens to be ill, feeding the chickens for about 20 days according to the group treatment related experimental group, and finally respectively using the ratio of the weights of spleen and thymus organs to the weights of animals and the serum superoxide dismutase (SOD) and Glutathione (GSH) values as indexes to prove the antioxidant effect of different treatment groups.

(1) Ratio of spleen to thymus organ weight to animal weight

Immune organ index (mg/g) =immune organ weight/living body weight.

(2) The specific operation steps of the determination of SOD are according to the instruction book of the SOD detection kit.

(3) GSH determination, specific operation steps are according to GSH detection kit instructions.

2. Experimental results and analysis:

(1) Results of ratio of spleen to thymus organ weight to animal weight

TABLE 5 results of ratio of spleen to thymus organ weight to animal weight

Grouping	A set of	Two groups of	Three groups of
				Spleen ratio size/%	1.13	1.16	1.10
Thymus ratio size/%	1.76	1.89	1.71

The immune organ index can be used as an important index for measuring the immunity of the poultry organism, and the increase of the weight of the immune organ indicates the enhancement of the immunity of the organism.

From Table 5, it can be seen that the ratio of the two groups is the greatest, and it can be seen that the two groups (fermentation broth added with the fungal elicitor) are better in index of spleen and thymus organs than the other two groups, which indirectly indicates that the fermentation broth obtained by adding the fungal elicitor has better effect of enhancing the immunity of the poultry.

(2) SOD and GSH value measurement results

TABLE 6 results of SOD and GSH values ratio

Grouping	A set of	Two groups of	Three groups of
				SOD(U/mg)	12.4	16.7	10.5
GSH(ng/mg)	86.9	92.1	73.7

As can be seen from table 6, the SOD values of the two groups were the largest; the maximum GSH value of two groups can be obtained, and two groups (fermentation liquor added with fungus elicitors) are better than other two groups in SOD and GSH cell factor indexes, so that the fermentation liquor obtained by adding the fungus elicitors has the functions of protecting cells and enhancing the oxidation resistance of organisms.

In conclusion, from the immune organ indexes (spleen and thymus) of the chickens and the measurement results of SOD and GSH values in serum, each index of the two experimental groups is superior to that of the other groups. This shows that the red-method yeast fermentation broth obtained by adding a mixture of marine red yeast cells and red yeast rice to a medium can improve the antioxidant capacity of broiler chickens.

The foregoing description of the preferred embodiments of the invention is not intended to be limiting, but rather is intended to cover all modifications, equivalents, alternatives, and improvements that fall within the spirit and scope of the invention.

Claims

1. A method for improving the astaxanthin yield of rhodozyma by using a fungal elicitor, comprising the steps of:

s1, preparing Phaffia rhodozyma seed liquid:

s2, preparing a fungus exciton:

s3, fermenting Phaffia rhodozyma:

s4, extracting astaxanthin:

2. The method for increasing the astaxanthin yield by using a fungal elicitor according to claim 1, wherein said fungal material is any one or two of yeast, mushroom, red yeast rice.

3. The method for increasing astaxanthin production by phaffia rhodozyma using a fungal elicitor according to claim 2, wherein the yeast is any one of rhodozyma marinum and rhodozyma mucilaginosa.

4. The method for increasing astaxanthin production by phaffia rhodozyma using a fungal elicitor according to claim 3, wherein the fungus is any of Coriolus versicolor, ganoderma lucidum, xuezhi, and black sesame.

5. The method according to claim 4, wherein the fungal material is any one of rhodotorula rubra, coriolus versicolor, red yeast rice, a mixture of a fermentation broth of rhodotorula rubra and a fermentation broth of coriolus versicolor, a mixture of coriolus versicolor and red yeast rice, and a mixture of rhodotorula rubra thallus and red yeast rice.

6. The method for increasing the astaxanthin production by a phaffia rhodozyma using a fungal elicitor according to claim 2, wherein the fungal material is a mixture of marine rhodozyma cells and red yeast rice.

7. The method for increasing the astaxanthin yield by using a fungal elicitor according to claim 2, wherein said step S2 comprises the specific steps of: respectively taking fungus raw materials, adding pure water, centrifuging at 3000-5000 rpm/min for 10-20 min, centrifuging for several times until the fungus raw materials are washed cleanly, taking precipitate, stirring, collecting crushed materials, freezing at-70 ℃ to-90 ℃ for 4-6 days, thawing in a water bath kettle at 50-70 ℃ for three times, repeatedly freezing and thawing, crushing the lyophilized materials, sieving with a 60-mesh sieve, and sterilizing for 20-50 min under high-pressure steam to obtain fungus elicitors.

8. The method for increasing astaxanthin production by phaffia rhodozyma according to claim 1, wherein said shake culture conditions in step S1 are: the temperature is 20-30 ℃, the rotating speed is 160-180 rpm/min, and the culture time is 3-5 days; the secondary shaking culture conditions in the step S1 are as follows: the temperature is 20-30 ℃, the rotating speed is 160-180 rpm/min, and the culture time is 2-4 days.

9. An astaxanthin formulation comprising an astaxanthin extract prepared according to the method of claim 1 that utilizes a fungal elicitor to increase the astaxanthin production by phaffia rhodozyma.

10. Use of an astaxanthin formulation according to claim 9 for feeding poultry.