阅读说明：本技术 一种含氯倍半萜衍生物及其制备和应用 (Chlorine-containing sesquiterpene derivative and preparation and application thereof ) 是由 季乃云 宋银平 于 2020-05-25 设计创作，主要内容包括：本发明涉及抑藻剂领域,具体地说是一种海藻内生真菌来源的含氯倍半萜衍生物及其制备方法和在抑藻方面的应用。具体结构式如(I)所示,其制备方法为将海藻内生真菌绿木霉(Trichoderma virens)RR-dl-6-8接种于真菌培养基中发酵培养,发酵产物经分离纯化后,即为式(I)所示的含氯倍半萜衍生物。本发明获得的含氯倍半萜衍生物经抑微藻活性实验得出化合物对微藻的半数抑制浓度可达0.83微克/毫升。<Image he="464" wi="459" file="DDA0002506452230000011.GIF" imgContent="drawing" imgFormat="GIF" orientation="portrait" inline="no"></Image>(The invention relates to the field of algae inhibiting agents, in particular to a chlorine-containing sesquiterpene derivative derived from algal endophytic fungi, a preparation method thereof and application thereof in algae inhibition. The preparation method comprises inoculating Trichoderma viride (Trichoderma virens) RR-dl-6-8 of marine algae endophytic fungus into a fungus culture medium for fermentation culture, and separating and purifying the fermentation product to obtain the chlorine-containing sesquiterpene derivative shown in formula (I). The chlorine-containing sesquiterpene derivative obtained by the invention inhibits microalgae activityExperiments show that the half inhibitory concentration of the compound on microalgae can reach 0.83 micrograms/milliliter.)

1. A chloro-sesquiterpene derivative, which is characterized in that: the structure of the chlorine-containing sesquiterpene derivative is shown as the formula (I)

2. A process for the preparation of chloro-sesquiterpene derivatives according to claim 1, wherein: inoculating Trichoderma viride (Trichoderma virens) RR-dl-6-8 into a fungus culture medium for fermentation culture, and purifying a fermentation product to obtain the chlorine-containing sesquiterpene derivative shown in the formula (I); the Trichoderma viride (Trichoderma virens) RR-dl-6-8 is preserved in the China Center for Type Culture Collection (CCTCC) in 2018, 6 months and 27 days, and the preservation number is CCTCC M2018406;

3. the process for the preparation of chlorinated sesquiterpene derivatives according to claim 2, comprising the specific steps of:

1) inoculating Trichoderma viride (Trichoderma virens) RR-dl-6-8 into a fungus culture medium, fermenting for 10-60 days, extracting by an organic solvent and concentrating to obtain a crude extract;

2) subjecting the crude extract obtained in the step 1) to silica gel column chromatography, performing gradient elution by using an organic solvent, collecting eluent, and detecting the eluent by thin layer chromatography;

3) collecting the eluted components in the step 2), and sequentially carrying out reversed phase silica gel column chromatography, gel column chromatography and preparative thin layer chromatography for separation and purification to obtain the chlorinated sesquiterpene derivative shown in the formula (I).

4. A process for the preparation of chlorinated sesquiterpene derivatives according to claim 3, wherein: the fungus culture medium in the step 1) is a rice solid culture medium, a jerusalem artichoke glucose liquid culture medium or a potato glucose liquid culture medium;

the organic solvent extract is one or more of ethyl acetate, petroleum ether, n-hexane, cyclohexane, dichloromethane, methanol, ethanol, propanol or isopropanol.

5. A process for the preparation of chlorinated sesquiterpene derivatives according to claim 3, wherein: the eluent in the step 2) is one or more of petroleum ether-ethyl acetate, petroleum ether-ethanol, petroleum ether-propanol, petroleum ether-isopropanol, dichloromethane-ethyl acetate, dichloromethane-methanol, dichloromethane-ethanol, dichloromethane-propanol and dichloromethane-isopropanol in a volume ratio of 50-0: 1.

6. A process for the preparation of chlorinated sesquiterpene derivatives according to claim 3, wherein: the reverse phase silica gel column chromatography eluent in the step 3) is water-methanol or water-ethanol with the volume ratio of 5-0: 1; the eluent of the gel column chromatography is dichloromethane-methanol or dichloromethane-ethanol with the volume ratio of 2-0: 1; the thin layer chromatography developing solvent is prepared from dichloromethane-methanol or petroleum ether-ethanol with the volume ratio of 30-0: 1.

7. Use of a chloro-sesquiterpene derivative according to claim 1, wherein: the chlorine-containing sesquiterpene derivative is applied to preparing an algistat for microalgae.

8. Use of chlorine-containing sesquiterpene derivatives according to claim 7 wherein: the algistat can inhibit one or more of red tide heterocurvula, robusta or prorocentrum donghaiense which are harmful to red tide.

Technical Field

The invention relates to the field of algae inhibiting agents, in particular to a chlorine-containing sesquiterpene derivative derived from algal endophytic fungi, a preparation method thereof and application thereof in algae inhibition.

Background

Worldwide, offshore red tide disasters are increasingly aggravated, and the development of marine fishery, offshore tourism and marine economy is seriously hindered. The main harmful forms of red tide mainly include damage to marine ecological balance, damage to marine fishery resources, damage to human health and influence on the development of marine tourism. There are about 300 species of microalgae in the ocean that can trigger red tide, and among them, there are about 80 species of toxic red tide algae. The scientific and effective prevention and treatment of red tide is one of the worldwide problems. The prevention and control strategy of red tide is to prevent the eutrophication of seawater, and also to inhibit the development of red tide organisms, namely red tide algae, and the method for inhibiting the development of the red tide algae is mainly chemical treatment. Although chemical agents such as copper sulfate, chlorine gas, etc. can rapidly and effectively kill or inhibit harmful algae or inhibit the growth and reproduction thereof, there are problems of harm to non-red tide organisms, drug residues, secondary pollution to the environment, etc.

Compared with the traditional chemical agents used in the treatment of the red tide, the marine organism-derived natural medicine has the advantages of strong pertinence, high safety, obvious activity and environmental friendliness, and provides a new idea for solving the problem of prevention and treatment of the red tide caused by red tide algae at present.

Disclosure of Invention

The invention aims to provide chlorine-containing sesquiterpene derivatives, and preparation and application thereof.

In order to achieve the purpose, the invention adopts the technical scheme that:

a chloro-sesquiterpene derivative has a structure shown in formula (I)

A method for preparing chlorine-containing sesquiterpene derivatives comprises inoculating Trichoderma viride (Trichoderma virens) RR-dl-6-8 in fungus culture medium, fermenting, and purifying to obtain chlorine-containing sesquiterpene derivatives represented by formula (I); the Trichoderma viride (Trichoderma virens) RR-dl-6-8 is preserved in the China Center for Type Culture Collection (CCTCC) in 2018, 6 months and 27 days, and the preservation number is CCTCC M2018406.

The method specifically comprises the following steps:

1) inoculating Trichoderma viride (Trichoderma virens) RR-dl-6-8 into a fungus culture medium, fermenting for 10-60 days, extracting by an organic solvent and concentrating to obtain a crude extract;

2) subjecting the crude extract obtained in the step 1) to silica gel column chromatography, performing gradient elution by using an organic solvent, collecting eluent, and detecting the eluent by thin layer chromatography;

3) collecting the eluted components in the step 2), and sequentially carrying out reversed phase silica gel column chromatography, gel column chromatography and preparative thin layer chromatography for separation and purification to obtain the chlorinated sesquiterpene derivative shown in the formula (I).

The fungus culture medium in the step 1) is a rice solid culture medium, a jerusalem artichoke glucose liquid culture medium or a potato glucose liquid culture medium.

The organic solvent extract is one or more of ethyl acetate, petroleum ether, n-hexane, cyclohexane, dichloromethane, methanol, ethanol, propanol or isopropanol.

The organic solvent in the step 2) is one or more of petroleum ether-ethyl acetate, petroleum ether-ethanol, petroleum ether-propanol, petroleum ether-isopropanol, dichloromethane-ethyl acetate, dichloromethane-methanol, dichloromethane-ethanol, dichloromethane-propanol and dichloromethane-isopropanol in a volume ratio of 50-0: 1.

The developing solvent for thin layer chromatography is one or more of petroleum ether-ethyl acetate, petroleum ether-ethanol and dichloromethane-methanol in a volume ratio of 100-0: 1.

The reverse phase silica gel column chromatography eluent in the step 3) is water-methanol or water-ethanol with the volume ratio of 5-0: 1; the eluent of the gel column chromatography is dichloromethane-methanol or dichloromethane-ethanol with the volume ratio of 2-0: 1; the thin layer chromatography developing solvent is prepared from dichloromethane-methanol or petroleum ether-ethanol with the volume ratio of 30-0: 1.

An application of chlorine-containing sesquiterpene derivative in preparing microalgae algistat.

The algistat can be used as one or more of red tide heterocurvulus, robusta or prorocentrum donghaiense causing red tide hazard.

The invention has the following advantages: the chlorine-containing sesquiterpene derivative is obtained by extracting and separating Trichoderma viride (Trichoderma virens) RR-dl-6-8 fermentation separated from marine red algae, namely, the pine node algae, and half inhibitory concentrations of the compound to red tide heterocurvulus, strong pre-ditch algae and east sea protozobium which can cause red tide are respectively 0.87 microgram/ml, 1.5 microgram/ml and 0.83 microgram/ml through a microalgae activity inhibition experiment.

The specific implementation mode is as follows:

the invention is further illustrated below with reference to the examples of embodiment.