Fusarium fujikuroi and its use in the simultaneous preparation of various group b and group a fumonisins

Abstract

The application discloses a fusarium fujikuroi and application thereof in synchronous preparation of multiple B group and A group fumonisins. Firstly, the application discloses a fusarium fujikuroi with a preservation number of CGMCC No.42734. Further, the application discloses application of the fusarium fujikuroi in synchronous preparation of multiple B group and A group fumonisins. The application first develops the WFU8601 strain capable of synchronously preparing FB1, FB2, FB3, FA1, FA2 and FA3, i.e., six kinds of fumonisins, and the fumonisins are rich in toxin types and stable in performance. In addition, the application can synchronously separate the six kinds of fumonisins monomers at one time through a two-step chromatography purification process combined with ELSD online monitoring and automatic fraction collection, the steps are simple and controllable, the six kinds of fumonisins have high recovery rate and purity, and the product quality can meet the research and development requirements of standard products, thereby providing a reliable material basis for fumonisins detection and toxicology research.

Description

Technical Field

[0001] This invention relates to the field of microbial technology. More specifically, it relates to a strain of Fusarium jirovecii and its application in the simultaneous preparation of various Group B and Group A fumonisins. Background Technology

[0002] Fumonisins are a class of secondary metabolites primarily produced by Fusarium microorganisms. Their structure is characterized by a long-chain, multi-hydroxyl carbon skeleton and two molecules of tricarboxylic acid, classifying them as lipid compounds. Since Bezuidenhout's initial identification in 1988, over 28 fumonisins have been discovered, classified into A, B, C, and P series based on structural differences. Group B fumonisins (FBs) are the most common and widely studied. Studies have shown that FBs exhibit various toxic effects, including neurotoxicity, hepatotoxicity, nephrotoxicity, enterotoxicity, immunotoxicity, reproductive toxicity, and combined toxicity. Their contamination is widespread globally, primarily found in food crops such as corn, rice, sorghum, and wheat, and their products, posing a serious threat to animal and human health through the food chain. Group A fumonisins (FAs), structurally similar to FBs, also exhibit cytotoxicity, with FA1 and FA2 contamination issues receiving increasing attention.

[0003] In research on fumonisins, including screening, formation mechanisms, metabolic patterns, hazard assessment, and detection techniques, high-purity, multi-variety fumonisin standards are indispensable basic materials. However, currently available fumonisin standards are expensive and of inconsistent quality, severely hindering in-depth research. Theoretically, preparative liquid chromatography (HPLC) can achieve efficient and high-purity preparation of fumonisin standards. However, because fumonisins themselves lack ultraviolet and fluorescence signals, existing detection techniques often require derivatization (a destructive process), resulting in very weak research on preparative HPLC techniques and few related literature reports. Research on group A fumonisins (FAs) is particularly limited. With a deeper understanding of the toxicity of FAs, developing a process that can simultaneously separate and prepare group B and group A fumonisins without derivatization has become an urgent technical problem to be solved in this field.

[0004] The preparation process of fumonisin standards mainly includes three steps: fermentation culture of toxin-producing strains, isolation and purification, and structural identification. Existing technologies have significant shortcomings in the following aspects: Regarding toxin-producing strains: Existing literature reports a limited number of fumonisin-producing strains, and most strains can only produce some types of group B fumonisins (such as FB1, FB2, and FB3), with very few strains capable of simultaneously producing group A fumonisins. The incomplete range of toxin-producing strains and low yields are among the main reasons for the lack of group A fumonisin standards and the insufficient supply of group B standards.

[0005] Regarding fermentation methods: existing fermentation methods are mainly divided into solid-state fermentation and liquid-state fermentation. Solid-state fermentation often uses substrates such as corn and rice, but it suffers from drawbacks such as long cultivation cycles, unstable yields, cumbersome operations (mostly using traditional methods such as conical flasks), and difficulty in scale-up. Although liquid-state fermentation offers better controllability, the yield of fumonisin is generally low under current process conditions. Neither method can meet the demand for stable, efficient, and large-scale preparation of various fumonisins.

[0006] Regarding separation and purification methods: existing purification technologies suffer from problems such as small throughput, cumbersome steps, and difficulty in balancing purity and yield. For example, a disclosed patent, CN 104931308 B, discloses a method for simultaneously preparing fumonisin B1, B2, and B3 standards. This method extracts the target toxins from toxin-contaminated rice and uses a cation exchange solid-phase extraction column combined with a two-step medium-pressure preparative liquid chromatography (CPLC) purification process. The limitations of this method are: small throughput from solid-phase extraction, long processing time in the two-step CPLC, and limited overall purification efficiency and scale; furthermore, it can only prepare three toxins from group B and cannot obtain group A fumonisins. CN 109503393 B discloses a method for preparing fumonisin B1 standards using high-speed countercurrent chromatography. This method extracts FB1 from rice cultures, separates it using a C18 column, and then purifies it using countercurrent chromatography. In this method, the FB1 content in the rice culture is approximately 400 mg / kg (approximately 368.76 mg of FB1 in 900 g of rice), but this method only targets the single FB1 toxin. CN 114317633 B discloses a liquid fermentation method for obtaining fungal toxin products by coupling nitrogen source, pH, and temperature induction. Through liquid fermentation and spray drying, group B fumonisins are obtained. The contents of fumonisins B1, B2, and B3 in the fermentation broth are 833 mg / L, 53.3 mg / L, and 21.4 mg / L, respectively. This method can only produce group B fumonisins and does not contain group A fumonisins. The spray drying of the fermentation broth has high energy consumption and time costs, and the fumonisin recovery rate is not mentioned. Subsequent chromatographic purification is still required.

[0007] Furthermore, the aforementioned three patents, as well as most current fumonisin preparation methods, employ offline detection methods (i.e., collecting fractions first, and then performing LC-MS / MS or TLC detection separately) because fumonisin lacks ultraviolet / fluorescence signals. This offline detection method is not only labor-intensive, time-consuming, and costly, but also cannot dynamically adjust and optimize the purification process in real time, greatly increasing the difficulty of obtaining high-purity toxin samples.

[0008] In summary, the existing technology has the following main drawbacks: there is a lack of toxin-producing strains capable of simultaneously producing high yields of multiple fumonisins from both Group B and Group A; there is a lack of fermentation and purification processes capable of simultaneously preparing Group B and Group A fumonisins (especially 6 or more); existing purification methods are inefficient, the purity and yield of the products are difficult to meet the requirements for the development of standard products, and they rely on offline detection, making process optimization impossible.

[0009] Therefore, developing a method for the simultaneous fermentation preparation, extraction, and purification of various B and A group fumonisins has significant scientific research value and practical application prospects. Summary of the Invention

[0010] The first objective of this invention is to provide Fusarium moniliforme capable of simultaneously producing multiple B and A fumonisins, specifically six B and A fumonisins.

[0011] The second objective of this invention is to provide the application of the aforementioned Fusarium moniliforme in the simultaneous preparation of various Group B and Group A fumonisins.

[0012] A third objective of this invention is to provide a method for simultaneously preparing various group B and group A fumonisins using the aforementioned Fusarium moniliforme.

[0013] To achieve the above objectives, the present invention adopts the following technical solution: This invention first provides a strain of *Fusarium oxysporum*, with accession number CGMCC No. 42734, and is classified as *Fusarium oxysporum*. Fusarium fujikuroi It was deposited on June 11, 2026 at the China General Microbiological Culture Collection Center (Address: No. 3, No. 1 Beichen West Road, Chaoyang District, Beijing).

[0014] This invention further provides the application of the above-mentioned Fusarium moniliforme in the simultaneous preparation of various Group B and Group A fumonisins.

[0015] In a specific embodiment of the present invention, the various B and A group fumonisins are FB1, FB2, FB3, FA1, FA2 and FA3.

[0016] The present invention further provides a method for simultaneously preparing various group B and group A fumonisins using the above-mentioned Fusarium oxysporum, comprising the following steps: Activation: Fusarium oxysporum was activated and cultured to obtain an activated bacterial solution; Solid-state fermentation: The activated bacterial solution is inoculated into a solid culture medium, water is added, and solid-state fermentation is carried out to obtain fermented material; Extraction: The fermentation material is dried, sieved, extracted, and filtered to obtain filtrate; Crude purification: The filtrate was concentrated, and the resulting extract was dissolved in 20% (v / v) methanol-water solution. Crude purification was performed using a medium-high pressure purification separation system (BUCHI Pure C-850 FlashPrep). The crude purification used a C18 Flash column with a sample loading volume of 0.5%~1% of the packing mass and a flow rate of 100~140 mL / min. Methanol-water solution was used as the mobile phase, and elution was performed according to the following procedure: first, impurities were eluted with 20%~40% (v / v) methanol-water solution, and then fumonisin was eluted with 60% methanol-water solution. The crude solution of Group B and Group A fumonisin was collected by online monitoring with an evaporative light scattering detector combined with an automatic fraction collector. Liquid phase separation and purification: The crude solutions of Group B and Group A fumonisins were dried, and the resulting powder was dissolved in a 20% (v / v) methanol-water solution containing 0.1% acetic acid. Separation was performed by high-performance liquid chromatography (Agela Technologies HP-Q-P050). The separation used a phenyl preparative column with a sample loading volume of 0.1%–0.3% of the packing mass, a flow rate of 20–40 mL / min, and a column temperature of 30–40 °C. The mobile phase was a methanol-water solution containing 0.1% acetic acid, and elution was performed according to the following linear gradient: 0–5 min, the methanol volume fraction increased linearly from 20% to 70%; 5–20 min, the methanol volume fraction increased linearly from 70% to 90%. The eluent of Group B and Group A fumonisins was collected by online monitoring with an evaporative light scattering detector combined with an automatic fraction collector. Drying treatment: The eluents of group B and group A fumonisin monomers were dried separately to obtain pure group B and group A fumonisin monomers; the group B and group A fumonisin monomers were FB1, FB2, FB3, FA1, FA2 and FA3, respectively.

[0017] In a specific embodiment of the present invention, the activation culture involves inoculating *Fusarium oxysporum* onto a PDA plate and incubating it statically at 25-30°C for 3-5 days to obtain mycelia; the mycelial sheets are then inoculated into PDB medium and cultured with shaking at 150-220 rpm at 25-30°C for 2-3 days; wherein each 1L of the PDA plate contains 10g of potato extract powder, 20g of glucose, and 20g of agar, and is sterilized at 115°C for 30 minutes; and each 1L of the PDB medium contains 10g of potato extract powder and 20g of glucose, and is sterilized at 115°C for 30 minutes.

[0018] In a preferred embodiment of the present invention, the activation culture involves inoculating Fusarium moniliforme onto a PDA plate and incubating it statically at 28°C for 3 days to obtain mycelia; the mycelial sheets are then inoculated into PDB medium and cultured at 28°C with shaking at 200 rpm for 2 days.

[0019] In a specific embodiment of the present invention, the solid culture medium is composed of 10%~30% corn husk and 70%~90% corn flour, and is prepared by sterilization at 115°C for 40 minutes; the mass-volume ratio of the solid culture medium to water is 5:2~5:3 (kg / L).

[0020] In a preferred embodiment of the present invention, the solid culture medium is composed of 20% corn husk and 80% corn flour; the mass-to-volume ratio of the solid culture medium to water is 5:3 (kg / L).

[0021] In a specific embodiment of the present invention, the solid-state fermentation bed is 3-5 cm thick, with a humidity of 70%-80%, a temperature of 20-30°C, and is cultured for 15-21 days.

[0022] In a preferred embodiment of the present invention, the solid-state fermentation bed is 3 cm thick, with a humidity of 80% and a temperature of 28°C, and is cultured for 15 days.

[0023] In a specific embodiment of the present invention, the drying and sieving are performed by drying at 60-80°C, pulverizing, and passing through a 20-50 mesh sieve; the extraction is performed by adding 60%-80% (v / v) methanol-water solution to the dried and sieved fermented material, and stirring at 100-300 rpm for 3-5 hours; the weight-volume ratio of the dried and sieved fermented material to the 60%-80% (v / v) methanol-water solution is 1:3-1:5 (kg / L).

[0024] In a preferred embodiment of the present invention, the drying and sieving are performed by drying at 80°C, pulverizing and passing through a 30-mesh sieve; the extraction is performed by adding an 80% (v / v) methanol-water solution to the dried and sieved fermented material and stirring at 200 rpm for 3 hours; the weight-to-volume ratio of the dried and sieved fermented material to the 80% (v / v) methanol-water solution is 1:3 (kg / L).

[0025] In a specific embodiment of the present invention, the filtrate concentration is performed under reduced pressure at 40-50°C and 60-150 mbar.

[0026] In a preferred embodiment of the present invention, the filtrate concentration is performed under reduced pressure at 45°C and 100 mbar.

[0027] In a specific embodiment of the present invention, the loading amount of the crude purified C18 Flash column is 0.5% of the packing mass, the flow rate is 120 mL / min, and the mobile phase is methanol-water solution. The elution is performed according to the following procedure: first, impurities are eluted with 40% (v / v) methanol-water solution, and then fumonisin is eluted with 60% methanol-water solution.

[0028] In a specific embodiment of the present invention, the crude fumonisin mixed solution is dried by rotary evaporation at 40-50°C.

[0029] In a preferred embodiment of the present invention, the crude fumonisin mixed solution is dried by rotary evaporation at 45°C.

[0030] In a specific embodiment of the present invention, the sample loading amount of the liquid phase separation and purification phenyl preparation column is 0.1% of the mass of the packing material, the flow rate is 20 mL / min, and the column temperature is 30 °C.

[0031] In a specific embodiment of the present invention, the parameters of the drift tube of the evaporative light scattering detector are set as follows: split ratio 1 / 30 to 1 / 15, drift tube temperature 60 to 80°C, and carrier gas flow rate 2 to 3 L / min.

[0032] In a preferred embodiment of the present invention, the parameters of the drift tube of the evaporative light scattering detector are: split ratio 1 / 30, drift tube temperature 80°C, and carrier gas flow rate 2 L / min.

[0033] In a specific embodiment of the present invention, the eluent is dried by concentrated under reduced pressure at 40-50°C and 60-150 mba, followed by freeze-drying until dry.

[0034] In a preferred embodiment of the present invention, the eluent is dried by concentration under reduced pressure at 45°C and 100 mba, followed by freeze-drying until dry.

[0035] The beneficial effects of this invention are as follows: The WFU8601 strain developed in this invention can simultaneously produce six fumonisins from both Group B (FB1, FB2, and FB3) and Group A (FA1, FA2, and FA3). It produces a wide variety of toxins with stable performance, providing a reliable strain source for the simultaneous preparation of multiple fumonisin monomers.

[0036] This invention establishes a method for the simultaneous preparation of six fumonisins using strain WFU8601. Through a two-step chromatographic purification process, combined with online ELSD monitoring and automatic fraction collection, pure monomers of six fumonisins, namely FB1, FB2, FB3, FA1, FA2 and FA3, can be obtained simultaneously in one step. The process steps are simple and the process is controllable.

[0037] The method of this invention produces six fumonisins with high yield and good purity, with yields ≥70% and purity ≥95%, meeting the quality requirements for the development of standard products and providing a reliable material basis for fumonisin detection and toxicological research. Attached Figure Description

[0038] The specific embodiments of the present invention will be described in further detail below with reference to the accompanying drawings.

[0039] Figure 1 Phylogenetic tree of strain WFU8601 based on ITS sequence.

[0040] Figure 2 This is a colony diagram of strain WFU8601.

[0041] Figure 3 The chromatograms show the separation of fumonisins FB1, FB2, FB3, FA1, FA2, and FA3.

[0042] Figure 4 Mass spectra of fumonisins FB1, FB2, FB3, FA1, FA2, and FA3.

[0043] Figure 5A Fumonisin FB1 1 H-NMR spectrum.

[0044] Figure 5B Fumonisin FB2 1 H-NMR spectrum.

[0045] Figure 5C Fumonisin FB3 1 H-NMR spectrum.

[0046] Figure 5D Fumonisin FA1 1 H-NMR spectrum.

[0047] Figure 5E Fumonisin FA2 1 H-NMR spectrum.

[0048] Figure 5F Fumonisin FA3 1 H-NMR spectrum.

[0049] Figure 6A Fumonisin FB1 13 C-NMR spectrum.

[0050] Figure 6B Fumonisin FB2 13 C-NMR spectrum.

[0051] Figure 6C Fumonisin FB3 13 C-NMR spectrum.

[0052] Figure 6D Fumonisin FA1 13 C-NMR spectrum.

[0053] Figure 6E Fumonisin FA2 13C-NMR spectrum.

[0054] Figure 6F Fumonisin FA3 13 C-NMR spectrum. Detailed Implementation

[0055] To more clearly illustrate the present invention, the following description, in conjunction with preferred embodiments and accompanying drawings, further explains the invention. Similar components in the drawings are indicated by the same reference numerals. Those skilled in the art should understand that the specific description below is illustrative rather than restrictive and should not be construed as limiting the scope of protection of the present invention.

[0056] Example 1: Fumonisin-producing strains Fusarium fujikuroi WFU8601 Isolation, Screening and Identification Strain WFU8601 was collected and isolated from maize kernels in a field in Xiangyang City, Hubei Province, and identified by ITS sequence ( Figure 1 (as shown) and colony morphology ( Figure 2 As shown), based on literature comparison, the strain was identified as Fusarium fusiforme (as shown). Fusarium fujikuroi ).

[0057] Strain WFU8601 was deposited on June 11, 2026, at the China General Microbiological Culture Collection Center (CGMCC, No. 3, No. 1 Beichen West Road, Chaoyang District, Beijing, 100101, China), with accession number CGMCC No. 42734, and classified as Fusarium oxysporum. Fusarium fujikuroi The following is abbreviated as Fusarium fujikuroi WFU8601.

[0058] Example 2 Using strains Fusarium fujikuroi WFU8601 also produces fumonisins (FB1, FB2, FB3, FA1, FA2, FA3). I. Preparation of Fumonisins (FB1, FB2, FB3, FA1, FA2, FA3) 1) Activation strain Fusarium fujikuroi WFU8601 was inoculated onto PDA plates (each 1L of medium contained 10g potato extract, 20g glucose, and 20g agar, prepared by sterilization at 115℃ for 30min) and incubated statically at 28℃ for 3 days to obtain mycelia; 6mm diameter mycelial sheets were obtained by punching holes in Oxford cups, and 3 sheets were inoculated into 1L of PDB medium (each 1L of medium contained 10g potato extract and 20g glucose, prepared by sterilization at 115℃ for 30min) and incubated at 28℃ with shaking at 200rpm for 2 days to obtain activated bacterial solution.

[0059] II) Solid-state fermentation Take 200 mL of activated bacterial solution and inoculate it into 1 kg of solid culture medium (the solid culture medium consists of 20% corn husk and 80% corn flour, sterilized at 115℃ for 40 min). Add 600 mL of sterile water and stir well to obtain the inoculated culture medium. Place the inoculated culture medium in a solid fermentation bed (20 cm × 40 cm) in an incubator with a bed thickness of 3 cm, humidity of 80%, and temperature of 28℃ for 15 days to obtain the fermented material.

[0060] III) Extraction After the fermentation material is dried at 80℃, it is crushed and passed through a 30-mesh sieve. The material is weighed after sieving, and fumonisin is extracted by adding 80% methanol-water solution at a material-to-liquid ratio of 1:3 (weight-to-volume ratio). The mixture is stirred at 200 rpm for 3 hours, filtered, and filtrate and residue are obtained. This process is repeated 3 times, and 3 filtrates are combined.

[0061] IV) Crude Purification The filtrate was concentrated under reduced pressure at 45°C and 100 mbar. The resulting extract was dissolved in a 20% (v / v) methanol-water solution and subjected to a first-step crude purification using a medium-high pressure purification system (BUCHI Pure C-850 FlashPrep) to rapidly enrich fumonisin and remove most impurities. Purification was performed using a C18 Flash column (packing material: YMC-ODS-AQ-HG, 12nm, S-50μm, packing mass 300g) at a flow rate of 120 mL / min, with a sample loading volume of 0.5% of the packing material mass. Elution was performed using a methanol-water solution as the mobile phase, following this procedure: first, impurities were eluted with a 40% (v / v) methanol-water solution, followed by fumonisin elution with a 60% methanol-water solution. Online monitoring was conducted using an evaporative light scattering detector (ELSD) (split ratio 1 / 30, drift tube temperature 80°C, carrier gas flow rate 2 L / min), combined with an automated fraction collector, to collect a mixed solution of crude Group B and Group A fumonisins. V) Liquid phase separation and purification A mixed solution of crude group B and group A fumonisins was rotary evaporated to dryness at 45°C. The resulting dry powder was dissolved in a 20% (v / v) methanol-water solution (containing 0.1% acetic acid) and used as the injection sample for the second purification step. Separation was performed using high-performance liquid chromatography (Agela Technologies HP-Q-P050). The preparative column was a phenyl preparative column (Welch-Phenyl, 5 μm, 21.2 × 250 mm), with a sample loading of 50 mg (0.1% of the packing mass), a flow rate of 20 mL / min, and a column temperature of 30°C. The mobile phase was a methanol-water solution containing 0.1% acetic acid, eluted with the following gradient: 20%–70% (v / v) methanol (0–5 min), 70%–90% (v / v) methanol (5–20 min). Online monitoring was performed using an evaporative light scattering detector (ELSD) (parameters set to a split ratio of 1 / 30, drift tube temperature of 80℃, and carrier gas flow rate of 2 L / min). Combined with an automated fraction collector, the eluents of each fumonisin monomer (FB1, FB2, FB3, FA1, FA2, FA3) were collected separately. Further processing yielded monomers with a purity exceeding 95%. Real-time online monitoring results of the chromatographic peaks of each monomer are shown below. Figure 3 .

[0062] vi) Drying treatment Each monomer eluent was concentrated to 100 mL under reduced pressure at 45 °C and 100 mba, and then lyophilized to dryness to obtain pure FB1, FB2, FB3, FA1, FA2 and FA3, respectively.

[0063] II. Recovery Rate (Yield) Calculation Yield = m 单体 / m 浸膏 *100%, where m 单体 This refers to the quality of pure FB1, FB2, FB3, FA1, FA2, and FA3; m 浸膏 The mass of each monomer in the extract is specified in GB5009.240-2023, and the mass of each monomer in the extract is determined by the second method of high performance liquid chromatography-tandem mass spectrometry.

[0064] Ultimately, the yields of pure FB1, FB2, FB3, FA1, FA2, and FA3 were all ≥70%.

[0065] III. Purity Testing The pure fumonisins FB1, FB2, FB3, FA1, FA2, and FA3 were detected by UPLC Q-TOF MS (6530) to obtain TIC chromatograms. The peak areas were normalized and calculated as follows: Purity = Peak area of ​​each monomer / (Total TIC peak area of ​​sample group - Total TIC peak area of ​​blank group) * 100%; where the total TIC peak area of ​​sample group refers to the sum of the peak areas of all ions in the TIC chromatograms of each sample; the total TIC peak area of ​​blank group refers to the sum of the peak areas of all ions in the blank group with only solvent added.

[0066] Ultimately, the purity of FB1, FB2, FB3, FA1, FA2, and FA3 was ≥95%.

[0067] IV. Verification 1) Obtaining and Results of High-Resolution Mass Spectrometry The pure fumonisins FB1, FB2, FB3, FA1, FA2, and FA3 were analyzed by Agilent UPLC-QTOF MS (6530), and the mass spectra of each monomer were obtained. The results are as follows. Figure 4 As shown, the results indicate that the mass spectrometry data (m / z) of the six Fumonis species are consistent with their molecular weights, and there are no other impurity signals, proving that the products are pure monomers of their respective species.

[0068] (ii) Obtaining and Results of NMR Spectra The pure fumonisins FB1, FB2, FB3, FA1, FA2, and FA3 were analyzed using a Bruker Advance NEO 600MHz nuclear magnetic resonance spectrometer to obtain the purity of each monomer. 1 H-NMR spectrum ( Figures 5A-5F )and 13 C-NMR spectrum ( Figures 6A-6F Spectral analysis showed that all relevant hydrogen and carbon signals of the six fumonisins were clearly distinguishable, and their chemical shifts ( 1 H and 13 C) The coupling constants of the hydrogen spectrum and the 1H spectrum are consistent with those reported in the literature, and no obvious impurity peaks were observed, thus confirming that each product is a single pure product.

[0069] Obviously, the above embodiments of the present invention are merely examples for clearly illustrating the present invention, and are not intended to limit the implementation of the present invention. For those skilled in the art, other variations or modifications can be made based on the above description. It is impossible to exhaustively list all the implementation methods here. All obvious variations or modifications derived from the technical solutions of the present invention are still within the protection scope of the present invention.

Claims

1. A strain of Fusarium oxysporum, characterized in that, The *Fusarium moniliforme* species described has the accession number CGMCC No. 42734 and is classified as *Fusarium moniliforme*. Fusarium fujikuroi .

2. The application of Fusarium oxysporum as described in claim 1 in the simultaneous preparation of various Group B and Group A fumonisins.

3. The application according to claim 2, characterized in that, The various B and A group fumonisins are FB1, FB2, FB3, FA1, FA2, and FA3.

4. The method for simultaneously preparing multiple group B and group A fumonisins using Fusarium oxysporum as described in claim 1, characterized in that, Includes the following steps: The *Fusarium moniliforme* was activated and cultured to obtain an activated bacterial solution; The activated bacterial solution was inoculated onto a solid culture medium, and water was added for solid-state fermentation to obtain fermented material. The fermented material is dried, sieved, extracted, and filtered to obtain a filtrate. The filtrate was concentrated, and the resulting extract was dissolved in a 20% methanol-water solution. The extract was then crudely purified using a medium-high pressure purification system. The crude purification employed a C18 Flash column with a loading volume of 0.5%–1% of the column mass and a flow rate of 100–140 mL / min. Methanol-water solution was used as the mobile phase, and elution was performed according to the following procedure: first, impurities were eluted with a 20%–40% methanol-water solution, then fumonisin was eluted with a 60% methanol-water solution. The mixture of crude Group B and Group A fumonisins was collected using an online evaporative light scattering detector combined with an automatic fraction collector. The crude solutions of Group B and Group A fumonisins were dried, and the resulting powder was dissolved in a 20% methanol-water solution containing 0.1% acetic acid. Separation was performed by high-performance liquid chromatography (HPLC) using a phenyl preparative column. The sample loading volume was 0.1%–0.3% of the packing material mass, the flow rate was 20–40 mL / min, and the column temperature was 30–40 °C. The mobile phase was a methanol-water solution containing 0.1% acetic acid, and elution was performed using the following linear gradient: 0–5 min, the methanol volume fraction increased linearly from 20% to 70%; 5–20 min, the methanol volume fraction increased linearly from 70% to 90%. The eluents of Group B and Group A fumonisins were collected using an online evaporative light scattering detector combined with an automatic fraction collector. The eluents of the group B and group A fumonisin monomers were dried separately to obtain pure group B and group A fumonisin monomers; the group B and group A fumonisin monomers were FB1, FB2, FB3, FA1, FA2 and FA3, respectively.

5. The method according to claim 4, characterized in that, The solid culture medium consists of 10% to 30% corn husks and 70% to 90% corn flour; the mass-to-volume ratio of the solid culture medium to water is 5:2 to 5:

3.

6. The method according to claim 4, characterized in that, The solid-state fermentation bed is 3-5 cm thick, with a humidity of 70%-80% and a temperature of 20-30℃, and is cultured for 15-21 days.

7. The method according to claim 4, characterized in that, The parameters of the evaporative light scattering detector are set as follows: split ratio 1 / 30~1 / 15, drift tube temperature 60~80℃, and carrier gas flow rate 2~3 L / min.

8. The method according to claim 4, characterized in that, The activation culture involves inoculating Fusarium moniliforme onto a PDA plate and incubating it statically at 25-30°C for 3-5 days to obtain mycelia; then inoculating the mycelial sheets onto PDB medium and incubating them with shaking at 150-220 rpm for 2-3 days at 25-30°C.

9. The method according to claim 4, characterized in that, The drying and sieving process involves drying at 60-80℃, followed by pulverizing and sieving through a 20-50 mesh sieve. Preferably, the extraction is carried out by adding 60%~80% methanol-water solution to the dried and sieved fermented material, and stirring at 100~300 rpm for 3~5 hours. More preferably, the weight-to-volume ratio of the dried and sieved fermented material to the 60%~80% methanol-water solution is 1:3~1:

5.

10. The method according to claim 4, characterized in that, The filtrate concentration is performed under reduced pressure at 40-50°C and 60-150 mbar. Preferably, the crude fumonisin mixed solution is dried by rotary evaporation at 40-50°C. More preferably, the eluent is dried by concentrated under reduced pressure at 40-50°C and 60-150 mba, followed by freeze-drying until dry.

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