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Polypeptin and preparation and application thereof

A technology of polypeptide bacteriocin and L-type, applied in the field of polypeptide bacterin and its preparation and application, can solve the problems of high residue, refractory degradation, high toxicity, etc., and achieve the effect of good safety and remarkable control effect.

Inactive Publication Date: 2011-02-02
ZHEJIANG UNIV +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, many chemical pesticides have the characteristics of high toxicity, high residue, and refractory degradation. With the widespread use of these pesticides, environmental pollution is becoming more and more serious, and the ecological balance is destroyed; the health of humans and livestock is also greatly threatened.
In addition, with the continuous emergence of drug-resistant bacteria and pests, the control efficiency of chemical pesticides has gradually declined. These problems have attracted people's attention, prompting people to look for a new control strategy that can replace chemical pesticides.

Method used

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  • Polypeptin and preparation and application thereof
  • Polypeptin and preparation and application thereof
  • Polypeptin and preparation and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0033] Example 1: The new compound of formula (I) can be produced by fermenting P.elgii NBRC 100335 according to the following conditions.

[0034] 1. Inoculate the strain NBRC 100335 on the slant of nutrient agar medium for activation, and culture it at 30°C for 1-2 years;

[0035] 2. Put the slant seeds into 50mL seed culture medium in a 250mL Erlenmeyer flask, and culture at 30°C and 200rpm for 24 hours, which is the first-grade seed solution;

[0036] 3. Put the first-grade seed solution into 200mL seed medium in a 500mL Erlenmeyer flask, and shake and culture at 30°C and 200rpm for 24h to obtain the second-grade seed solution;

[0037] 4. Add 4L of fermentation medium to a fermenter with a total volume of 7L, add 0.1% polyether defoamer GPE, sterilize under high pressure steam at 115°C for 60min, cool to 30°C, and put 200ml of secondary seed liquid into the fermenter (5% inoculum size, v / v), the initial rotation is 450 rpm, the dissolved oxygen is kept above 30%, and the...

Embodiment 2

[0041] Embodiment 2: the separation and purification of formula (I) novel compound

[0042] The fermentation broth obtained in Example 1 can be separated and purified according to the following steps to obtain the pure product of the new compound of formula (I) provided by the present invention.

[0043] 1. Transfer the fermentation liquid to a centrifuge bottle, centrifuge at 6000rpm for 15 minutes, remove the bacteria, and obtain the fermentation supernatant;

[0044] 2. Add an equal volume of n-butanol to the supernatant, shake and extract, collect the extract, and repeat the raffinate with an equal volume of n-butanol.

[0045] 3. The two extracts were combined and concentrated to dryness in vacuo at 65° C. with a rotary evaporator to obtain a n-butanol extract concentrate.

[0046] 4. The n-butanol extract concentrate was resuspended in water, passed through an MCI GEL CHP20P (Mitsubishi Chemical) column, and then eluted with 20%, 70% and 100% methanol respectively, and ...

Embodiment 3

[0048] Embodiment 3: antibacterial activity assay

[0049] 1. Determination of the minimum inhibitory concentration of bacteria

[0050] 1) The new compound of formula (I) with a concentration of 200 μg / ml was prepared with Mueller Hinton broth medium, and doubled: 200, 100, 50, 25, 12.5, 6.25, 3.12, 1.56, 0.78, 0.39 μg / ml ml.

[0051] 2) Add 50 μl of the new compound solution of formula (I) at different concentrations (200-0.39 μg / m1) after serial dilution to the 1st to 10th wells of the 96-well polystyrene plate, and add 50 μl of Mueller Hinton to the 11th well Broth medium (0.2% beef extract powder, 1.75% acid hydrolyzed casein, 0.15% starch) was used as a growth control, and 100 μl of Mueller Hinton broth medium was added to the 12th well as a negative control. Positive controls were polymyxin B and vancomycin.

[0052] 3) After culturing the indicator bacteria for 24 hours, dilute to 10 with Mueller Hinton broth medium 6 cells / ml, add 50 μl of diluted bacterial soluti...

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Abstract

The invention provides novel polypeptin having potential value of biological control and medical applications, and a preparation method and application thereof. The structure of the polypeptin is shown as a formula (I). The invention has the advantages of providing the polypeptin which has the toxicity lower than that of polymyxin B and has the potential value of biological control and medical applications, and the preparation method and the application thereof. In vitro experiments show that the compound has the obvious effect of inhibiting all the tested gram-positive bacterium, gram-negative bacterium and fungi. Disk-culturing experiments show that the compound has certain effect of preventing and treating various fungal plant diseases and particularly has obvious effects of preventing and treating rice sheath blight.

Description

(1) Technical field [0001] The present invention relates to a novel polypeptide bacterin and its preparation and application. (2) Background technology [0002] Globally, plant diseases caused by pathogenic bacteria and pests are one of the main causes of agricultural losses. For half a century, people have mainly used chemical pesticides to prevent and control these diseases, and have achieved remarkable results. However, many chemical pesticides have the characteristics of high toxicity, high residue, and refractory degradation. With the widespread use of these pesticides, environmental pollution has become increasingly serious, ecological balance has been destroyed, and the health of humans and livestock has also been greatly threatened. In addition, with the continuous emergence of drug-resistant bacteria and pests, the control efficiency of chemical pesticides has gradually declined. These problems have attracted people's attention, prompting people to look for a new c...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A01P3/00C07K1/20C12P21/02C07K7/06C07K1/14C12R1/01A01N37/46
Inventor 钱朝东李欧沈小波白骅吴雪昌杜加秋林建秋
Owner ZHEJIANG UNIV
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