A strain producing phenolic hydroxylated compounds and its use

Abstract

The application belongs to the technical field of microorganisms, and provides a strain for producing phenolic hydroxyl compounds and application thereof, wherein the strain is JW-24-8, which has been preserved in the China General Microbiological Culture Collection Center (CGMCC) on July 20, 2026, and the address of the CGMCC is No. 1, Beichen West Road, Chaoyang District, Beijing, and the preservation number is CGMCC No. 42846. The application provides a strain JW-24-8 for producing phenolic hydroxyl compounds, wherein the strain has excellent characteristics for producing phenolic hydroxyl compounds, and it is found through fingerprint spectrum analysis of a fermentation crude extract obtained by fermentation of the strain that a compound with multiple phenolic hydroxyl groups is the peak compound at 23 min, the yield is large, and there is no obvious impurity peak around the compound peak, which indicates that the strain provided in the application is more likely to obtain phenolic hydroxyl compounds with high purity and high yield compared with previous microbial strains for producing phenolic hydroxyl compounds.

Description

Technical Field

[0001] This invention belongs to the field of microbial technology, and in particular relates to a strain that produces phenolic hydroxyl compounds and its applications. Background Technology

[0002] Phenolic hydroxyl compounds possess both excellent physicochemical properties and diverse biological applications: the phenolic hydroxyl group endows the molecules with amphiphilic solubility, facilitating extraction, purification, and translocation and absorption within plants. Furthermore, the hydroxyl site is easily chemically modified to optimize activity. These substances can inhibit plant pathogens such as citrus canker through multi-target action, are less likely to induce antibiotic resistance, and can also induce disease resistance, antioxidant capacity, and stress tolerance in fruit trees. Phenolic hydroxyl compounds are mostly derived from natural microbial secondary metabolites, are easily degraded in the environment, have good ecological compatibility, and exhibit low toxicity to humans and animals, meeting the requirements for green pesticide development. Simultaneously, they can be produced on a large scale through microbial fermentation, making resource acquisition convenient and possessing broad development prospects in the field of agricultural plant protection. Summary of the Invention

[0003] The purpose of this invention is to provide a strain that produces phenolic hydroxyl compounds and its applications, thereby addressing the problems mentioned in the background section.

[0004] To achieve the above objectives, the present invention provides the following technical solution: On the one hand, the present invention provides a strain that produces phenolic hydroxyl compounds, the strain being JW-24-8, which was deposited on July 20, 2026 at the China General Microbiological Culture Collection Center, located at No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing, with accession number CGMCC No. 42846.

[0005] On the other hand, the present invention provides the application of a strain that produces phenolic hydroxyl compounds in the production of phenolic hydroxyl compounds.

[0006] On the other hand, the present invention provides the application of a strain that produces phenolic hydroxyl compounds in the preparation of an anti-citrus canker drug.

[0007] Compared with the prior art, the specific beneficial effects of the present invention are as follows: This invention provides a strain JW-24-8 that can produce phenolic hydroxyl compounds. This strain has excellent characteristics in producing phenolic hydroxyl compounds. Fingerprint analysis of the crude extract obtained by fermentation using this strain revealed that the eluting compound at 23 min is a compound containing multiple phenolic hydroxyl groups, and the yield is high. At the same time, there are no obvious impurity peaks around the compound peak. This indicates that the strain provided by this invention is more likely to produce phenolic hydroxyl compounds with high purity and high yield compared with previous microbial strains that produce phenolic hydroxyl compounds. Attached Figure Description

[0008] Figure 1 The fingerprint spectrum of the crude extract provided in the embodiments of the present invention; Figure 2 The 1H NMR spectrum of the peak components provided in the embodiments of the present invention; Figure 3 The carbon NMR spectrum of the peak components provided in the embodiments of the present invention; Figure 4 The high-resolution mass spectrum of the peak components provided in the embodiments of the present invention. Detailed Implementation

[0009] To make the objectives, technical solutions, and advantages of this invention clearer, the invention will be further described in detail below with reference to the accompanying drawings and embodiments. It should be understood that the specific embodiments described herein are merely illustrative and not intended to limit the invention.

[0010] The application of a strain that produces compounds containing phenolic hydroxyl groups in the production of compounds containing phenolic hydroxyl groups includes the following steps: S1. The strain was inoculated into the fermentation medium and fermented at 25-30℃ for 30 days to obtain the fermentation product. S2. The fermented material was soaked in ethyl acetate, and the ethyl acetate extract obtained by stirring and separation was then subjected to vacuum distillation to recover the solvent and obtain a compound containing phenolic hydroxyl groups. Furthermore, the strain was inoculated into the fermentation medium and cultured statically at 25–30°C for 30 days to obtain the fermentation product; Furthermore, before inoculating the strain into the fermentation medium, the strain is further subjected to a large-scale culture. The culture medium used for the large-scale culture is preferably a PDA medium with water as the solvent. The PDA medium is prepared as follows: 200g of peeled and diced potatoes are boiled in water for 20 minutes, filtered through gauze to obtain the extract, 20g of glucose and 18g of agar are added to the extract, and the volume is adjusted to 1L with water. The mixture is then steam-sterilized at 121°C under high temperature and high pressure for 30 minutes. The temperature of the large-scale culture is preferably 28°C, the environment of the large-scale culture is preferably a biochemical incubator, and the culture time is 3 days. Furthermore, the fermentation medium is preferably contained in a 1L Erlenmeyer flask, and the fermentation medium is 1L of water containing the following components by weight: 20g mannitol, 20g maltose, 10g glucose, 10g monosodium glutamate, 0.5g KH2PO4, 0.3g MgSO4·7H2O, 3g yeast extract, 1g corn steep liquor, pH 6.5; Furthermore, the fermentation temperature is preferably 28℃, and the fermentation time is preferably 30 days; static culture is preferred during fermentation, and the fermentation environment is preferably a biochemical incubator, which is beneficial for maintaining a constant temperature. Furthermore, the soaking time is preferably 10-14 hours, more preferably 12 hours, as soaking helps to dissolve penicillic acid in the fermentation product into ethyl acetate; the soaking temperature is preferably 20-27°C, more preferably 22-25°C; stirring extraction helps to enhance the complete dissolution of phenolic hydroxyl compounds into ethyl acetate; Furthermore, the pressure of the vacuum distillation is preferably -0.1 to 0 MPa relative pressure; the water bath temperature of the vacuum distillation is preferably 20 to 25°C; there are no special restrictions on the method of vacuum distillation, and any vacuum distillation method well known in the art can be used. Furthermore, the recovered solvent yielded a crude penicillic acid extract. Fingerprint analysis was performed on the crude penicillic acid extract, and the concentrations of the mobile phase analyzed were as follows: 10% methanol aqueous solution (volume concentration) for 0–5 min; 5–45 min, volume concentration of 10%–100% methanol aqueous solution.

[0011] The following detailed description, in conjunction with embodiments, illustrates a strain that produces phenolic hydroxyl compounds and its application in agricultural anti-ulcer disease, but these descriptions should not be construed as limiting the scope of protection of this invention.

[0012] The specific implementation of the present invention will be described in detail below with reference to specific embodiments.

[0013] Example 1, Screening of strains: Moss was collected from the Jinggangshan area of ​​Ji'an City for subsequent isolation. The samples were ground and inoculated onto PDA medium for isolation and culture at 28°C. After morphological identification, the strain obtained was identified as JW-24-8, which was deposited on July 20, 2026, at the China General Microbiological Culture Collection Center (CGMCC), located at No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing, with accession number CGMCC No. 42846.

[0014] Example 2: Application of strain JW-24-8 in the production of compounds containing phenolic hydroxyl groups, specifically including the following steps: The JW-24-8 strain isolated in Example 1 was inoculated onto a microbial culture medium (PDA medium) to obtain the strain. A culture medium containing the following components was prepared as the fermentation medium: 6g mannitol, 6g maltose, 3g glucose, 3g monosodium glutamate, 0.15g KH2PO4, 0.09g MgSO4·7H2O, 0.9g yeast extract, 0.3g corn steep liquor, and 300mL water were added to each 1L Erlenmeyer flask. The medium was then autoclaved to obtain the fermentation medium. The JW-24-8 strain was inoculated into 120 Erlenmeyer flasks and incubated at a constant temperature of 28℃ for 30 days to obtain the fermentation product. The fermentation broth was subjected to solid-liquid separation. The bacterial culture was collected and extracted with ethyl acetate. The obtained ethyl acetate phase was subjected to vacuum distillation to recover the solvent (evaporator: water bath temperature 35℃, circulating water vacuum pump to maintain the apparatus under vacuum, vacuum gauge reading approximately -0.1MPa relative pressure). Finally, 3.0g of crude extract was obtained. The crude extract was subjected to fingerprint analysis under the following conditions: 0–5 min: 10% methanol aqueous solution (v / v); 5–45 min: 100% methanol aqueous solution (v / v); 45–50 min: pure methanol. The fingerprint chromatogram of the crude extract is shown below. Figure 1 The compound that elutes at 23 minutes was detected to contain phenolic hydroxyl compounds.

[0015] The crude extract was passed through a normal-phase column with silica gel as the stationary phase and a mobile phase of A:ethyl acetate = 1:1 (where A is petroleum ether:ethyl acetate = 1:1). The active fraction was obtained, and the resulting mixture was subjected to high-pressure liquid chromatography (HPLC) to obtain a purified extract. A 21% methanol-water solution was used as the mobile phase. The extract elutes at 23 minutes, and the peak components were collected for NMR analysis. The resulting 1H NMR spectrum is shown below. Figure 2 As shown, the carbon spectrum is as follows Figure 3 As shown, the mass spectrum is as follows Figure 4 As shown, it has been identified as a phenolic hydroxyl compound, and its structural formula is shown below: ; According to ultraviolet light detection, the purity of the product extracted by this invention is above 99.5%.

[0016] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, and improvements made within the spirit and principles of the present invention should be included within the protection scope of the present invention.

Claims

1. A strain that produces phenolic hydroxyl compounds, characterized in that, The strain in question is JW-24-8, which was deposited on July 20, 2026, at the China General Microbiological Culture Collection Center (CGMCC), located at No. 3, Courtyard 1, Beichen West Road, Chaoyang District, Beijing, with accession number CGMCC No. 42846.

2. The use of a strain for producing phenolic hydroxyl compounds as described in claim 1 in the production of phenolic hydroxyl compounds.

3. The application according to claim 2, characterized in that, Includes the following steps: S1. Inoculate strain JW-24-8 into liquid fermentation medium and allow it to ferment statically to obtain fermentation product; S2. The fermented material was soaked in ethyl acetate, and the ethyl acetate extract obtained by extraction and separation was then subjected to vacuum distillation to recover the solvent, yielding a compound containing phenolic hydroxyl groups.

4. The application according to claim 3, characterized in that, In S1, the liquid fermentation medium contains the following components by weight in 1L of water: 20g mannitol, 20g maltose, 10g glucose, 10g monosodium glutamate, 0.5g KH2PO4, 0.3g MgSO4·7H2O, 3g yeast extract, and 1g corn steep liquor. The pH of the liquid fermentation medium is 6.

5.

5. The application according to claim 3, characterized in that, In S1, the fermentation culture temperature is 28℃ and the time is 30 days.

6. The application according to claim 3, characterized in that, In S2, the pressure of the vacuum distillation is a relative pressure of -0.1 to 0 MPa, and the water bath temperature for vacuum distillation is 25°C.

7. The application according to claim 3, characterized in that, In step S2, the process of recovering the solvent further includes purifying the recovered crude extract; the purification method includes the following steps: S21. The recovered crude extract is separated by vacuum silica gel normal phase column chromatography, and the fraction is collected. S22. The fraction is separated by high-pressure liquid phase to obtain a compound containing phenolic hydroxyl groups.

8. The application according to claim 7, characterized in that, In S21, the stationary phase of the vacuum silica gel normal phase column is 200-300 mesh column chromatography silica gel, and the mobile phase is petroleum ether, ethyl acetate, and methanol, eluted in a gradient manner.

9. The application according to claim 7, characterized in that, In S22, the mobile phase used for high-pressure liquid phase separation is a methanol-water solution with a volume concentration of 21%.

10. The use of a strain that produces phenolic hydroxyl compounds as described in claim 1 in the preparation of an anti-citrus canker drug.

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