CN112514896B

CN112514896B - Rhodiola sachalinensis wettable powder and application

Info

Publication number: CN112514896B
Application number: CN202011417680.9A
Authority: CN
Inventors: 马贵龙; 高新馨; 赵成爱; 袁守超; 张婷婷
Original assignee: Jilin Agricultural University
Current assignee: Jilin Agricultural University
Priority date: 2020-12-07
Filing date: 2020-12-07
Publication date: 2022-02-01
Anticipated expiration: 2040-12-07
Also published as: CN112514896A

Abstract

The invention discloses rhodiola sachalinensis wettable powder which comprises the following components in percentage by mass: 45-48% of rhodiola aqueous extract, 45-48% of carrier, 2-3% of wetting agent and 3-5% of dispersing agent. When the carrier is white carbon black (400 meshes), the wetting agent is 2 percent of sodium dodecyl benzene sulfonate and the dispersing agent is 3 percent of sodium polynaphthalenesulfonate, the wetting time of the wettable powder is 27.88s, the suspension rate is 74.44 percent, the bacteriostasis rate of the rhodiola sachalinensis wettable powder to rhizoctonia solani is 85.06 percent when the concentration is 1mg/mL, and the bacteriostasis rate to the rhodiola sachalinensis damping-off bacteria is 92.41 percent when the concentration is 0.05 mg/mL.

Description

Rhodiola sachalinensis wettable powder and application

Technical Field

The invention belongs to the technical field of natural product extraction, and particularly relates to rhodiola sachalinensis wettable powder and application thereof.

Background

Rhodiola sachalinensis (A. boreale)Rhodiolasachalinensis) Also called rhodiola sachalinensis, is a perennial herb with extremely strong vitality. Has more obvious effects in the resistance tests of anti-anoxia, anti-radiation, anti-fatigue, anti-virus and the like. The rhodiola sachalinensis is mainly produced in the counties of Changbai mountain area in Jilin province, Heilongjiang province, Shanzhi, Hai forest, Ning ' an and the like, wherein the Changbai mountain area in Jilin province is firstly planted in the last 80 th century, has a large development area and is known as ' the village of rhodiola rosea ', and the mountain area in the northeast three province is planted in a large area at present. The rhodiola sachalinensis mainly contains salidroside and tyrosol, and further contains starch, protein, fat, tannin, flavonoid compounds, trace volatile oil, and trace elements such as iron, zinc, tin, molybdenum, manganese, etc. Can be used for treating sexual hypofunction, diabetes, and hypotension.

Ginseng (A, B)Panax ginseng C.A.Meyer) Is a perennial root plant of Araliaceae, is produced in northeast, Korea, Japan and the east of Russia in China, and is known as the king of all grass. At present, more than 40 ginseng diseases are proved at home and abroad in totalAbout 25 diseases have been found in the planting process of the ginseng. The disease is classified into 2 types, i.e., invasive disease and non-invasive disease, according to whether the disease is infectious or not. The infectious diseases comprise sclerotinia, gray mold, epidemic disease, damping off, black spot, damping off, rust rot, etc.; non-invasive diseases include root splits, red skin disease, etc. Dividing the diseases into seedling diseases and growth diseases according to different growth stages of ginseng. The diseases in the seedling stage mainly comprise 2 damping-off diseases and damping-off diseases; the long-term diseases of the ginseng mainly comprise gray mold, black spot, bacterial soft rot, root rot, red skin disease and the like. Epidemic disease, rust rot, sclerotinia, etc. may appear in each stage of ginseng growth and development.

Most of the ginseng farmers can control ginseng diseases by using medicaments such as fenaminosulf, mancozeb, carbendazim, ethephon, procymidone, metalaxyl manganese zinc, pulex and the like, and the control is mainly based on chemical medicaments, and the generation of drug-resistant strains is caused by excessive spraying times in the using process; the chemical pesticide is used as a main prevention and treatment means for ginseng diseases and plays an important role in the production process of ginseng. Compared with other control modes, the pesticide control operation is relatively simple, and the pesticide control method has the characteristics of large control scale, quick pesticide effect and the like. However, in the process of disease control in part of ginseng farmers and ginseng factories, the problems of unreasonable pesticide use, single pesticide use and the like exist, which also causes the phenomena of ginseng planting yield reduction, drug resistance enhancement and the like.

The biological control of ginseng diseases mainly uses bacillus subtilis and trichoderma harzianum which have obvious control effects on seedling damping off and root rot of sown ginseng and transplanted ginseng. In recent years, more and more scientific researchers are actively involved in the screening work of the biocontrol bacteria agent, and many biocontrol bacteria with bacteriostatic activity are continuously discovered, but a plurality of practical problems still exist when the biocontrol bacteria agent is really applied to field production. The conditions required by fermentation production, preparation processing, storage and application of the biological agent are very strict and greatly change along with different floras, whether the viable bacteria preparation can exert effective inhibition on pathogenic bacteria and the degree of action of the viable bacteria preparation is greatly influenced by environmental factors, and some bio-control strains with obvious bacteriostatic effect in a laboratory show the phenomena of bacteriostatic activity reduction, difficult colonization, strain decline and the like in the actual application process, so that the bio-control strains temporarily do not have commercial objective conditions, which requires scientific researchers to develop efficient and stable living bio-control agents and simultaneously consider that antibiotics, antimicrobial proteins and other antibacterial substances generated by the bio-control bacteria can be prepared into microbial pesticides, so that the bio-control agents can be better served for agricultural production.

At present, along with the gradual expansion of the production scale of pollution-free agricultural products and the increasing severity of the problems of drug resistance, pesticide residue and the like caused by the use of chemical pesticides, the searching and the use of nontoxic novel plant source pesticides with high efficiency are more and more valued by people. The plant source pesticide rhodiola sachalinensis extract for inhibiting ginseng pathogenic bacteria has not been reported.

Disclosure of Invention

The invention provides a rhodiola sachalinensis wettable powder for solving the problems.

The rhodiola sachalinensis wettable powder comprises the following components in percentage by mass: 45 to 48 percent of rhodiola aqueous extract,

45% -48% of carrier, 2% -3% of wetting agent and 3% -5% of dispersing agent;

the rhodiola aqueous extract is prepared by the following method:

drying radix Rhodiolae, soaking in water, filtering, collecting filtrate and residue, decocting the residue in water, filtering, collecting filtrate, mixing the filtrates, concentrating, and freeze drying to obtain radix Rhodiolae water extract;

the carrier is white carbon black (400 meshes);

the ratio of the carrier dosage to the rhodiola aqueous extract is 1:1

The wetting agent is sodium dodecyl benzene sulfonate;

the mass percentage of the wetting agent is 2 percent;

the dispersant is sodium polynaphthalenesulfonate;

the mass percent of the dispersant is 3 percent;

the rhodiola sachalinensis wettable powder is applied to inhibition of rhizoctonia solani and ginseng damping-off.

The rhodiola sachalinensis extract is prepared by the following method:

drying radix Rhodiolae, soaking in water, filtering, collecting filtrate and residue, decocting the residue in water, filtering, collecting filtrate, mixing the filtrates, concentrating, and freeze drying to obtain radix Rhodiolae extract;

the soaking time is 4-6 days;

the decocting time is 15-25 min;

the rhodiola sachalinensis extract is dynamically adsorbed and eluted by macroporous resin, the rhodiola sachalinensis extract is firstly eluted by water, the eluent is discarded, then the rhodiola sachalinensis extract is eluted by 10% ethanol solution, the eluent is collected, and the solvent is removed by evaporation;

the macroporous resin is D101 type macroporous resin;

the rhodiola sachalinensis extract is applied to inhibiting ginseng damping-off bacteria and ginseng rhizoctonia solani.

The invention provides rhodiola sachalinensis wettable powder which comprises the following components in percentage by mass: 45-48% of rhodiola aqueous extract, 45-48% of carrier, 2-3% of wetting agent and 3-5% of dispersing agent. When the carrier is white carbon black (400 meshes), the wetting agent is 2 percent of sodium dodecyl benzene sulfonate and the dispersing agent is 3 percent of sodium polynaphthalenesulfonate, the wetting time of the wettable powder is 27.88s, the suspension rate is 74.44 percent, the bacteriostasis rate of the rhodiola sachalinensis wettable powder to rhizoctonia solani is 85.06 percent when the concentration is 1mg/mL, and the bacteriostasis rate to the rhodiola sachalinensis damping-off bacteria is 92.41 percent when the concentration is 0.05 mg/mL.

Drawings

FIG. 1 shows the bacteriostatic action of ethanol eluates with different concentrations on rhizoctonia solani;

FIG. 2 shows the bacteriostatic action of ethanol eluate with different concentrations on damping-off of ginseng;

FIG. 3 shows the effect of different carriers of rhodiola sachalinensis aqueous extract on rhizoctonia solani;

FIG. 4 shows the effect of different carriers on damping off of ginseng in rhodiola sachalinensis aqueous extracts.

Detailed Description

Example 1 test materials

Test materials: radix Rhodiolae dry root. The producing area: the product is purchased from Lanxin specialty trade company, Inc. of Baishan City, Jilin province.

Test standards and reagents: salidroside (standard), tyrosol (standard), rosavil (standard), and kaempferol (standard). All 4 standards were purchased from Chenguang Biotech, Inc., Baoji City.

Reagent to be tested: 50% carbendazim wettable powder (Jiangsu Lanfeng biochemistry industry Co., Ltd.), 30% methyl-hymexazol water aqua (Zhejiang Hebei science and technology Co., Ltd.), and 2.5% fludioxonil suspended seed coating agent (Xiongdatong crop protection Co., Ltd.).

Test strains: ginseng rhizoctonia solani bacterium (A)Rhizoctonia solani) Ginseng damping-off bacteria (1)Pythium debaryanum) Ginseng root rot pathogen (A)F. oxysporum）Ginseng black spot pathogenAlternaria panax) Ginseng botrytis cinerea (A.Meyer)Botrytis cinerea) Ginseng sclerotinia sclerotiorum (A) and (B)Sclerotinia sclerotiorum) Ginseng rust rot (A), (B), (C)Cylindrocarpon destructans) Ginseng phytophthora genus (A)Phytophthora cactorum) Botrytis cinerea (A), (B), (C), (B), (C), (B), (C), (B), (C), (B) a), (B) a) and a)Botrytis cinerea) Cucumber fusarium wilt bacteria (Fusarium oxysporum (Schl.) f.sp ) Sclerotinia sclerotiorum (A) and (B) of cucumberSclerotinia sclerotiorum) (supplied by the plant pathology laboratory).

Carrier: kaolin, calcium carbonate, white carbon black, bentonite, diatomite and attapulgite.

Wetting agent: sodium dodecyl sulfate, sodium dodecyl benzene sulfonate and nekal BX (made in China).

Dispersing agent: calcium lignosulfonate, sodium polynaphthalene formaldehyde sulfonate and disodium methylenedinaphthalene sulfonate.

Test medium: the preparation method of the PDA culture medium comprises the steps of 200g of potatoes, 20g of glucose, 18-20g of agar and constant volume of 1L of distilled water.

Example 2 preparation of aqueous extract of rhodiola sachalinensis

Weighing 20g of rhodiola sachalinensis dry root, placing in a proper container, adding 100 times of distilled water, soaking for 5 days. Filtering with non-woven fabrics after soaking, and respectively collecting filtrate and filter residue. Placing the filter residue in a stainless steel pot, adding 30 times of distilled water into the pot, and decocting for 20 min. Filtering with non-woven fabric after decocting, and collecting rhodiola sachalinensis filtrate. Mixing the filtrate soaked in distilled water with the decoction filtrate, concentrating with a rotary evaporator, and stopping operation when the volume of the concentrated solution is about 10 mL. Freeze drying to obtain rhodiola sachalinensis aqueous extract, sealing and storing in brown reagent bottle, and placing in refrigerator at 4 deg.C for use.

Example 3 preparation of extract of rhodiola sachalinensis

Placing a certain volume of macroporous resin (D101) in a big beaker, slowly adding an ethanol solution with the volume concentration of 95% into the beaker until the liquid level of the ethanol solution exceeds macroporous resin particles, standing for about 12 hours, washing the ethanol solution until the ethanol solution is odorless, then loading the ethanol solution into a column after the ethanol solution is washed to two thirds of the column, preparing the rhodiola sachalinensis aqueous extract into the concentration of 5mg/mL for loading, wherein the loading amount is one third of the volume of the macroporous resin, standing for 12 hours after loading, washing the rhodiola sachalinensis aqueous extract by sequentially using distilled water, 10%, 20% and 30% … 80% ethanol, collecting different elution components, concentrating and drying the eluate into powder, and sequentially numbering a: water eluate, b: 10% ethanol eluate, c:20% ethanol eluate, d: i, 30% ethanol eluate.: 80% ethanol eluate.

Example 4 bacteriostatic activity of rhodiola sachalinensis aqueous extract

1. Determination of antibacterial activity of rhodiola sachalinensis aqueous extract

The aqueous extract obtained in example 2 was prepared as a solution at a concentration of 15mg/mL for use. The growth rate method is adopted to determine the inhibiting effect of the rhodiola sachalinensis aqueous extract on 11 plant pathogenic bacteria. The cross method used in this test measures the colony diameter. The results are shown in table 1, when the concentration of the rhodiola sachalinensis aqueous extract is 15mg/mL, the rhodiola sachalinensis aqueous extract has different degrees of inhibition effects on 10 plant pathogenic bacteria, wherein the inhibition rate on the ginseng damping-off and the ginseng rhizoctonia solani can reach 100%, the inhibition rate on the ginseng root rot, the ginseng sclerotinia sclerotiorum and the cucumber fusarium wilt can reach more than 80%, the inhibition rate on the other 5 bacteria is less than 55%, and the rhodiola sachalinensis aqueous extract has no inhibition effect on the cucumber sclerotinia sclerotiorum.

The aqueous extract obtained in example 2 was prepared into a solution having a concentration of 2mg/mL or 3mg/mL for use. The growth rate method is adopted to determine the inhibition effect of the rhodiola sachalinensis aqueous extract on the ginseng damping-off mildew. The results are shown in Table 2, when the concentration of the rhodiola sachalinensis aqueous extract is 2mg/mL, the inhibition rate on the ginseng damping-off pathogen can reach 100%.

The aqueous extract obtained in example 3 was prepared as a solution at a concentration of 3mg/mL for use. The growth rate method is adopted to determine the inhibition effect of the rhodiola sachalinensis aqueous extract on rhizoctonia solani. The results are shown in Table 3, when the concentration of rhodiola sachalinensis aqueous extract is 3mg/mL, the bacteriostasis rate of the rhodiola sachalinensis aqueous extract on the rhizoctonia solani is 50%.

2. Inhibition effect of 4 monomer compound standard substances of rhodiola sachalinensis on 2 pathogenic bacteria of ginseng

The growth rate method is used for measuring the inhibiting effect of 4 monomer compound standard substances of the rhodiola sachalinensis on 2 pathogenic bacteria of the ginseng. The concentration of the 4 monomeric compound standard substances in a culture medium plate is 0.5mg/mL, and when the control group bacterial colony grows over the culture dish plate, the bacterial colony diameters of each test group and the control group are measured and the bacteriostasis rate is calculated. As shown in Table 4, the 4 monomeric compound standards have no bacteriostatic effect on ginseng rhizoctonia solani and ginseng damping-off.

TABLE 4 inhibition of 2 pathogens of ginseng by different rhodiola sachalinensis standards

Standard article	The bacteriostasis rate of the ginseng rhizoctonia solani is%	Bacteriostasis rate of ginseng damping-off pathogen%
			Salidroside
	0	0
			Kaempferol	0	0
Luoxinwei medicine	0	0
			Tyrosol	0	0

EXAMPLE 5 bacteriostatic test and virulence determination for each component of the refined extract

The eluate of example 3 a: water, b: respectively preparing a PDA culture medium plate containing 1mg/mL of each eluate by using 10% ethanol eluate, c, 20% ethanol eluate and i, and measuring the bacteriostatic activity of the PDA culture medium plate on the rhizoctonia solani and the ginseng damping-off germ by using a growth rate method. The other operation steps for determining the bacteriostasis are the same as the example 4, and the results are shown in figures 1 and 2, wherein the rhodiola sachalinensis aqueous extract 10% ethanol eluate, and the rhodiola sachalinensis aqueous extract 60% ethanol eluate and the rhodiola sachalinensis aqueous extract 70% ethanol eluate have the bacteriostasis on rhizoctonia solani. Wherein the 10% ethanol eluate has the best antibacterial effect, the antibacterial rate is more than 80%, and other 6 eluates have no antibacterial effect on Ginseng radix Rhizoctonia solani. 9 eluates obtained by separating the rhodiola sachalinensis aqueous extract have bacteriostasis effect on ginseng damping-off germs, wherein the 10% ethanol eluate has the best bacteriostasis effect, and the bacteriostasis rate reaches 100%.

PDA medium plates were prepared containing 10% ethanol eluate concentrations (2.5, 2.0, 1.5, 1.0, 0.5 mg/mL) and (0.5, 0.4, 0.3, 0.2, 0.1 mg/mL). Respectively carrying out toxicity determination on the ginseng rhizoctonia solani and the ginseng damping-off germ. From Table 5, the 10% ethanol eluate of rhodiola sachalinensis has an EC50 value of 0.6122mg/mL for rhizoctonia solani and an EC50 value of 0.2013mg/mL for damping-off. The EC50 value of the rhodiola sachalinensis 10% ethanol eluate for damping-off bacteria is smaller than the EC50 value of the rhizoctonia solani, which shows that the bacteriostatic activity of the rhodiola sachalinensis 10% ethanol eluate for damping-off bacteria has better effect on the ginseng than the rhizoctonia solani.

Example 6 preparation of wettable powders

1. Vector screening

2g of each carrier sample was accurately weighed into a small beaker, linseed oil was added dropwise and stirred with a glass rod until the sample began to clump together without breaking apart. The saturated oil absorption was calculated based on the amount of linseed oil consumed, taking care not to overdose the linseed oil. Repeat 3 times and average. Referring to GB/T5451-2001, the wetting time of the carrier is measured, the shorter the time, the better the wetting property, and the wetting time can not exceed 2 min. The measurement of the suspension percentage of the carrier refers to GB/T14825-2006, the higher the suspension percentage is, the better the dispersibility is, and the suspension percentage cannot be lower than 70%. The growth rate method is adopted to determine the bacteriostatic effect of the rhodiola sachalinensis aqueous extract added into each carrier on the rhizoctonia solani and the ginseng damping-off germ, and the optimal carrier is selected by comparing the factors of the carriers in the aspects of oil absorption rate, wetting time, suspension rate, cost and the like. Mixing the carrier and the rhodiola sachalinensis aqueous extract according to a ratio of 1:1, drying, and grinding into powder to obtain mother powder.

As shown in Table 6, the oil absorption of the carrier silica (40 mesh) was 6.60ml, the oil absorption of the next silica (400 mesh) was 5.01ml, and the oil absorption of diatomaceous earth was 3.35ml, and since 61.10% of the suspension of silica (400 mesh) was larger than 55.55% of silica (40 mesh), and the suspension of diatomaceous earth was 12.22%, and the wetting time of silica (400 mesh) was 15.64s, which was shorter than 49.66 s of silica (40 mesh) 32.56 s, silica was selected as the carrier. By adopting the hypha growth rate method, as shown in table 7 (summarized in fig. 3 and 4), white carbon black (400 mesh) has a better effect of inhibiting rhizoctonia solani and the like. In conclusion, white carbon black (400 meshes) is selected as a carrier for developing the rhodiola sachalinensis wettable powder.

2. Wetting agent screening

Mixing a wetting agent into the mother powder to prepare wettable powder, and determining the wetting time of the preparation according to GB/T5451-2001, wherein the shorter the time, the better the wettability; the measurement of the suspension rate of the preparation refers to GB/T14825-2006, and the higher the suspension rate is, the better the dispersibility is; and detecting the bacteriostasis effect on the ginseng rhizoctonia solani by a growth rate method, measuring the bacteriostasis diameter, and screening out the optimal wetting according to the result. Different contents of 1%, 2%, 3%, 4% and 5% are added, and the optimal addition amount of the wetting agent is selected.

As can be seen from Table 8, the wetting time of adding sodium dodecyl benzene sulfonate into the mother powder is the shortest, the wetting time is 18.59s and 20.30s respectively after the powder is pulled out, the bacteriostasis rates to the rhizoctonia solani are 78.48 percent and 45.82 percent respectively, and the bacteriostasis rates to the damping-off bacteria are 93.29 percent and 92.78 percent respectively. Sodium dodecylbenzenesulfonate was therefore chosen as wetting agent. As can be seen from Table 9, the suspension percentage can reach 71.34% when the content of the sodium dodecyl benzene sulfonate is 2%, the wetting time is 19.97s, and 2% -3% is selected as the optimal addition amount of the sodium dodecyl benzene sulfonate.

3. Dispersant screening

Mixing a dispersing agent into mother powder added with a wetting agent to prepare wettable powder, and determining the wetting time of the preparation with reference to GB/T5451-2001, wherein the shorter the time is, the better the wettability is; the measurement of the suspension rate of the preparation refers to GB/T14825-2006, and the higher the suspension rate is, the better the dispersibility is; and detecting the bacteriostasis effect on the ginseng rhizoctonia solani by a growth rate method, measuring the bacteriostasis diameter, and screening out the optimal dispersant according to the result. Different contents of 1%, 2%, 3%, 4% and 5% are added, and the optimal addition amount of the dispersing agent is selected.

As can be seen from Table 10, the wetting times after the addition of the 4 dispersants are not very different, wherein the wetting time of the sodium polynaphthalenesulfonate is the shortest, and the sodium polynaphthalenesulfonate is selected in combination with the results of the bacteriostasis test. As shown in Table 11, the difference in wetting time was not so large at the sodium polynaphthalenesulfonate contents of 3%, 4% and 5%, which were 27.88s, 27.78s and 27.16s, respectively. 3% -5% is selected as the optimal addition amount by combining the suspension rate and the bacteriostasis test.

When the concentration of the rhodiola sachalinensis wettable powder is 1mg/mL, the bacteriostasis rate of the rhodiola sachalinensis wettable powder to rhizoctonia solani is 85.06%, the bacteriostasis rate of a 30% metalaxyl-hymexazol water agent is 83.42%, the bacteriostasis rate of a 35% metalaxyl seed treatment dry powder agent is 63.82%, and the bacteriostasis rate of a 50% carbendazim wettable powder is 100%. The rhodiola sachalinensis wettable powder has a comparable bacteriostatic effect with a 30% metalaxyl-hymexazol water aqua, and the bacteriostatic rate is higher than that of 35% metalaxyl seed treatment dry powder.

When the concentration of the rhodiola sachalinensis wettable powder is 0.05mg/mL, the inhibition rate of the rhodiola sachalinensis wettable powder on the ginseng damping-off germs is 92.41%, under the same dosage, the inhibition rate of a 30% hymexazol-formosan aqueous solution is 90.51%, the inhibition rate of a 2.5% fludioxonil suspension seed coating agent is 72.15%, and the inhibition rate of a 50% carbendazim wettable powder is 1.03%. The rhodiola sachalinensis wettable powder has a comparable bacteriostatic effect with a 30% fine methyl hymexazol water aqua, and the bacteriostatic rate is far higher than that of 50% carbendazim wettable powder.

Claims

1. The rhodiola sachalinensis wettable powder is characterized by comprising the following components in percentage by mass: 45-48% of rhodiola aqueous extract, 45-48% of carrier, 2-3% of wetting agent and 3-5% of dispersing agent;

the carrier is 400-mesh white carbon black;

the ratio of the dosage of the carrier to the rhodiola aqueous extract is 1: 1;

the wetting agent is sodium dodecyl benzene sulfonate.

2. The rhodiola sachalinensis wettable powder of claim 1, wherein the aqueous extract of rhodiola sachalinensis is prepared by the following method:

soaking dried radix Rhodiolae in water, filtering, collecting filtrate and residue, decocting the residue in water, filtering, collecting filtrate, mixing the filtrates, concentrating, and freeze drying to obtain radix Rhodiolae water extract.

3. The rhodiola sachalinensis wettable powder according to claim 2, characterized in that: the mass percentage of the wetting agent is 2 percent.

4. The rhodiola sachalinensis wettable powder according to claim 3, characterized in that: the dispersant is sodium polynaphthalenesulfonate.

5. The rhodiola sachalinensis wettable powder according to claim 4, characterized in that: the mass percent of the dispersant is 3%.

6. The use of the rhodiola sachalinensis wettable powder of claim 1 for inhibiting rhizoctonia solani and ginseng damping-off.