CN105326824A - Application of palmitic acid in preparation of medicine for killing external parasites of aquatic animals - Google Patents

Abstract

The invention discloses application of palmitic acid in preparation of a medicine for killing external parasites of aquatic animals and further discloses a prevention and control agent containing palmitic acid and used for killing external parasites of aquatic animals. The palmitic acid is a novel pollution-free green fishery medicine, has low toxicity to fish, does not easily enable the parasites to producing medicine resistance, can efficiently kill dactylogyrus and is remarkable in effect. The production cost of the palmitic acid is low, a using method of the palmitic acid is simple, and the palmitic acid is convenient to popularize and apply.

Description

Palmic acid is preparing the application in killing aquatic animal vermin medicine

Technical field

The present invention relates to Palmic acid (C ₁₆h ₃₂o ₂) novelty teabag, particularly relate to a kind of Palmic acid and preparing the application in killing aquatic animal vermin medicine.

Background technology

Palmic acid [16 (alkane) acid, palmitic acid] is led a cow (Pharbitisnil) or mature seed---a kind of compound of extraction and isolation Semen Pharbitidis of Pharbitis purpurea (Pharbitispurpurea) from medicinal plants annual vine Convolvulaceae ipomoea plant, and its structural formula is:

The molecular formula of this compound is C ₁₆h ₃₂o ₂, molecular weight is 256.4, and white flaky solid is water insoluble, is slightly soluble in cold alcohol and petroleum ether, is dissolved in hot ethanol, EC etc.For the manufacture of chemical articles such as wax candle, soap, metallic soap, grease, synthetic detergent, softening agents, or the intermediate etc. of pharmaceutical synthesis.

Summary of the invention

In order to the shortcoming overcoming prior art is with not enough, a kind of Palmic acid is the object of the present invention is to provide to prepare application in killing aquatic animal vermin medicine.

Another object of the present invention is to provide a kind of killing aquatic animal external parasite preventing and treating agent.

Another object of the present invention is the preparation method providing above-mentioned killing aquatic animal external parasite preventing and treating agent.

Object of the present invention is achieved through the following technical solutions:

Palmic acid is preparing the application in killing aquatic animal vermin medicine.

Further, Palmic acid is preparing the application killed in Fish epizoa medicine.

Further, Palmic acid is preparing the application killed in Fish epizoa-dactylogyrider medicine.

Present invention also offers a kind of killing aquatic animal external parasite preventing and treating agent, be made up of the raw material of following masses percentage ratio: Palmic acid 1% ~ 20%, surfactant 5% ~ 10%, transdermal agent 1% ~ 4%, all the other are solvent, and the summation of above-mentioned raw materials is 100%;

Described surfactant is preferably tween 20;

Described transdermal agent is preferably water-solubleazone (XW-02), and name is called 1-orthododecyl nitrogen heterocyclo heptane-2-ketone, molecular formula C ₁₈h ₃₅nO;

Described solvent is preferably dehydrated alcohol.

The preferred solvent of the present invention is dehydrated alcohol, can also select other solvents well known to those skilled in the art, such as methanol.

The preparation method of described killing aquatic animal external parasite preventing and treating agent, comprises the following steps:

(1) Palmic acid, surfactant, transdermal agent, each raw material of solvent is taken, for subsequent use;

(2) Palmic acid and solvent are mixed to dissolve completely, obtain mixed solution;

(3) surfactant, transdermal agent are added in the mixed solution of step (2) gained stir, obtain killing aquatic animal external parasite preventing and treating agent.

The using method of killing aquatic animal external parasite preventing and treating agent of the present invention is simple, directly by 12 ~ 20mL/m ³dosage splash and can effectively to prevent and treat or killing aquatic animal (Fish) vermin, particularly dactylogyrider (Dactylogyrusspp.) in pond.

The present invention, relative to prior art, has following advantage and effect:

(1) the present invention has excavated novelty teabag to known compound Palmic acid, has opened up new opplication field.

(2) Palmic acid is the novel fisheries drug of a kind of nuisanceless green, low to toxicity in fish, not easily cause parasite produce drug resistance, efficiently can kill dactylogyrider, Be very effective.

(3) production cost of Palmic acid is low, and using method is simple, easy to utilize.

Detailed description of the invention

Below in conjunction with embodiment, the present invention is described in further detail, but embodiments of the present invention are not limited thereto.

In order to understand essence of the present invention better, below will with Palmic acid to fish acute toxicity experiment, pharmacological testing, the pharmacological testing of control agent and result illustrate its novelty teabag in killing aquatic animal vermin medicine.

Bad gill dactylogyrider (Dactylogyrusvastator) document " 29 kinds of natural plant extracts are to the research [J] of dactylogyrider killing action. fresh water fishery, 2006,03:3-8. " in open.

Embodiment 1 Palmic acid is to Brachydanio rerio acute toxicity test

1 materials and methods

1.1 material

1.1.1 test sample

Testing Palmic acid purity used is more than 95.0%, and being prepared by this laboratory provides.Accurately take 1.0g Palmic acid, fully dissolve with dehydrated alcohol (purchased from Tianjin chemical reagent factory), standardize solution is in 100mL volumetric flask, and being made into concentration is 10.0mg/mL, and-4 DEG C of Refrigerator stores are for subsequent use.

1.1.2 experimental animal

Test Brachydanio rerio used (Daniorerio) to be purchased from and to view and admire fishing ground, require that body constitution is healthy and strong, specification is homogeneous, average weight 0.45 ± 0.06g.In laboratory aquarium, support more than 7d temporarily after buying back, throw something and feed every day 1 ~ 2 time, the feedstuff of throwing something and feeding is eaten up and is limited in 10min.Treat Brachydanio rerio physical health, well-grown, after colony is basicly stable, carry out toxicity test.

1.1.3 test apparatus

One of ALC-1100.2 percentage electronic balance, Beijing Sai Duolisi instrument system company limited; KQ-500E type ultrasonic cleaner; 5 ~ 10 μ L, 20 ~ 200 μ L pipettor: VoluMate, Germany; Automatic temperature control heating rod, Zhoushan Zhejiang fishing gear factory produces; Plastic tub (volume is 8L) etc.

1.2 test method

1.2.1 experimental condition

Adopt hydrostatic formula test method(s), every basin adds the tap water 6L of abundant aeration, pH7.0 ~ 7.2, and controlling water temperature is 25 ± 1 DEG C, and first add and to add test after test medicinal liquid fully mixes again with fish, 10 tails put in a suitable place to breed by every basin.After putting fish in a suitable place to breed, require to keep dissolved oxygen in water at more than 5mg/L.In order to keep constant drug level, change water dosing again every 24h.

1.2.2 the setting of trial drug concentration

First, repeatedly carry out trial test, determine the approximate range of formal test liquor strength, namely test the highest liquor strength (4.5mg/L) that death does not occur in the fish minimum liquor strength (9.0mg/L) of being all poisoned to death in 24h and 96h.Then on trial test basis, within the scope of liquor strength 4.5 ~ 9.0mg/L, insert 14 concentration tests groups by log concentration equal difference (tolerance d=0.3), carry out formal test, under observing each concentration, test the situation of being poisoned to death of fish.

In the death condition of duration of test observed and recorded test at any time fish, as found, ichthyism is dead, immediately should pull out, in order to avoid affect water quality, affect result of the test.Judge that the method for fish death is the caudal peduncle portion that (opercular movement stopping) touching fish with Glass rod or tweezers after fish ceases breathing, if fish body does not produce any stress within 3min, can death be judged.

Duration of test statistics tests the death condition of fish in 24h, 48h, 72h and 96h under each drug level.Duration of test should be kept quite, and avoids any interference to test fish as far as possible.When the mortality rate of matched group is less than 5%, in each time period, the mortality rate of test fish can correct; If the mortality rate of matched group is greater than 5%, then should correct, updating formula adopts Abbott formula; If matched group mortality rate reaches more than 20%, then, after searching reason, re-start toxicity test.

Abbott formula: p=(p '-C)/(1-C);

P: corrected mortality, i.e. sheerly drug-induced mortality rate;

P ': test group mortality rate, i.e. natural cause and drug-induced mortality rate;

C: matched group mortality rate, the i.e. mortality rate that causes of natural cause.

1.2.3 Palmic acid minimum lethal concentration scope, median lethal concentration (LC ₅₀)

Observed and recorded is for the death condition of examination fish in 24h, 48h, 72h and 96h, and test fish each time period starts to occur that dead least concentration and next concentration are the minimum lethal concentration scope of this time period, and calculates fatality rate.Utilize return law of the straight line, with the linear regression equation y=a+bx (log concentration of x-drug level; The probability unit that y-mortality rate converts to; A, b are respectively intercept and the slope of straight line.), obtain a and b, the computing formula of a and b is:

$a=\frac{{\mathrm{\Σx}}^2\mathrm{\Σ}y-\mathrm{\Σ}x\mathrm{\Σ}\left(xy\right)}{{\mathrm{k\Σx}}^2-{\left(\mathrm{\Σ}x\right)}^2}$

$b=\frac{k\mathrm{\Σ}\left(xy\right)-\mathrm{\Σ}x\mathrm{\Σ}y}{{\mathrm{k\Σx}}^2-{\left(\mathrm{\Σ}x\right)}^2}$

K: the group number of drug level (mortality rate be 0 and 100% concentration do not count)

After obtaining a, b, just can determine regression equation, and then bring the probability unit y=5.0 of 50% mortality rate into equation, obtain x, get its antilogarithm, just draw the LC in this time ₅₀, and calculate its fiducial limit of 95%, formula is:

In formula: $s=\frac{1}{b}$

N: for examination animal number (mortality rate be 0 and 100% concentration do not count)

X:LC ₅₀the logarithm of value

1.2.4 the calculating of Palmic acid safe concentration

According to Turubell formulae discovery safe concentration:

2 results and analysis

2.1 Palmic acid median lethal concentration (LC ₅₀)

2.1.1 each experimental concentration of Palmic acid in different time to Brachydanio rerio intoxicating situation

The Brachydanio rerio death condition of each trial drug concentration in 24h, 48h, 96h is in table 1.In the high concentration test group that drug level is 9.0mg/L, clearly, after medication there is Novel presentation in about 2h to the reaction of test ichthyism, head emerges by Brachydanio rerio, is then slow in action, and occurs that side is swum, start in the near future to occur paralysis, disequilibrium, abdominal part is dead upward.Be in the low concentration test group of 4.5mg/L in concentration, Brachydanio rerio performance is quieter, is quick on the draw to external world, movable normal.

Table 1 Palmic acid is to the result of being poisoned to death of Brachydanio rerio

2.1.2 minimum lethal concentration scope, median lethal concentration (LC ₅₀)

The death condition of statistics variable concentrations test medicinal liquid in 12h, 24h, 48h and 96h, the results are shown in Table 2.

Each experimental concentration is got its logarithm, obtain its log concentration, and its probit that the percentage rate of each time period test ichthyism death is tabled look-up to obtain, the probit of the drug level logarithm of each time period and the mortality rate of correspondence thereof is substituted into formula, obtain regression equation, the median lethal concentration (LC of each time period can be calculated ₅₀), and its LC ₅₀95% fiducial limit.The results are shown in Table 3.

Table 2 Palmic acid is to the acute toxicity result of Brachydanio rerio

The LC of table 3 each time period ₅₀and the fiducial limit of 95%

As shown in Table 3,

In 12h, the regression equation of gained is: y=-35.363x+34.617, LC ₅₀=7.058mg/L, 95% be crediblely limited to 6.770 ~ 7.360mg/L;

In 24h, the regression equation of gained is: y=-36.025x+35.035, LC ₅₀=6.982mg/L, 95% be crediblely limited to 6.702 ~ 7.276mg/L;

In 48h, the regression equation of gained is: y=-35.752x+34.435, LC ₅₀=6.794mg/L, 95% be crediblely limited to 6.506 ~ 7.089mg/L;

In 96h, the regression equation of gained is: y=-35.752x+34.435, LC ₅₀=6.459mg/L, 95% be crediblely limited to 6.157 ~ 6.750mg/L.

2.1.3 safe concentration result of calculation

The safe concentration of Palmic acid to Brachydanio rerio is gone out according to Turbell formulae discovery.

From result display, the safe concentration of Palmic acid to Brachydanio rerio is 1.930mg/L, but in reality, concentration is in 4.5mg/L situation, and 96h duration of test occurs without death; Meanwhile, when 4.8mg/L, 48h does not occur that fish is dead, is 1.4mg/L (LC in conjunction with the effective dose killing dactylogyrider ₅₀), consider, Palmic acid has good application and popularization value as pesticide for fish.

Embodiment 2 Palmic acid is to dactylogyrider killing effect

1 materials and methods

1.1 experimental animal

Host's model is Carassius auratus (Carassiusauratus), is purchased from pet fish market, Guangzhou, body weight 5.0 ± 0.5g; Parasite is bad gill dactylogyrider (Dactylogyrusvastator), and this laboratory from abstraction and purification the Carassius auratus gill, and to go down to posterity conservation with Carassius auratus.Bad gill dactylogyrider (Dactylogyrusvastator) document " 29 kinds of natural plant extracts are to the research [J] of dactylogyrider killing action. fresh water fishery, 2006,03:3-8. " in open.

1.2 trial drug

Testing Palmic acid purity used is more than 90.0%, is that Fructus Arctii (Arctiumlappa) is separated obtains through extraction, multistage silica gel column chromatography.

1.3 test method

Use the plastic tub that volume is 8L, contain into abundant aeration tap water 5L, use automatic temperature control heating rod to control water temperature 25 ± 1 DEG C, pH7.0 ~ 7.5.During on-test, first by random samples 10 tails are inspected at random to the Carassius auratus material having infected dactylogyrider, when parasite mean parasitized quantity and infection rate reach test requirements document (50 ~ 60/sheet gill and 100%), throw in respectively in each basin and infect Carassius auratus 10 tail.Then adding each flow point in aforementioned each separating step respectively in water body according to testing the concentration set up, stirring.The film-making of depletion fish holobranch, microscopy during 48h, observe various sample killing effect to dactylogyrider under variable concentrations, fish mortality rate when statistics optimum effective concentration, the highest killing rate, ichthyism concentration and poisoning concentration.Test repetition 3 times, and establish matched group.

Get in the setting parasite killing time and observe for the film-making of examination fish holobranch, microscopy and add up dactylogyrider from gill portion dropping situations, if do not come off, whether deadly observe, around polypide cephalic gland, mouthpart does not shrink person at 1 ~ 2min, show for death, if without worm or worm dead, gill portion all determines that killing rate is 100%; If partial exfoliation, add up every tail fish gill portion survival parasite quantity, calculating mean value, and contrast with matched group, calculate killing rate, evaluate insecticidal effect with this.

2.1 kill dactylogyrider effect

2.1.1 the insecticidal activity assay of Palmic acid activated monomer

Purity is that 90.0% Palmic acid measures it and kills dactylogyrider activity in table 4.

Table 4 Palmic acid 48h kills activity to dactylogyrider

As shown in Table 4, when the concentration of Palmic acid is from 3.0mg/L, 100% is reached to the killing rate of dactylogyrider; And when concentration reaches 8.0mg/L, start to show certain toxicity to fish, the mortality rate of this macrura reevesii is 10%.

Essence for a better understanding of the present invention.The specific embodiment provided below in conjunction with inventor, to describe Palmic acid killing aquatic animal external parasite preventing and treating agent preparation method in detail, is below good embodiment, but is not limited to these embodiments.

Embodiment 3

Take after Palmic acid 1.0% and solvent dehydrated alcohol 86.0% mix to and dissolve completely according to mass percent, add surface active agent tween-2010.0% and transdermal agent water-solubleazone 3.0%, abundant mix homogeneously, obtains killing aquatic animal external parasite preventing and treating agent.

Embodiment 4

Killing aquatic animal external parasite preventing and treating agent, mass percent is: Palmic acid 4.0%, surface active agent tween-208.0%, transdermal agent water-solubleazone 2.0%, solvent dehydrated alcohol 86.0%.Preparation method is with embodiment 3.

Embodiment 5

Killing aquatic animal external parasite preventing and treating agent, mass percent is: Palmic acid 8.0%, surface active agent tween-2010.0%, transdermal agent water-solubleazone 2.0%, solvent dehydrated alcohol 80.0%.Preparation method is with embodiment 3.

Embodiment 6

Killing aquatic animal external parasite preventing and treating agent, mass percent is: Palmic acid 12.0%, surface active agent tween-207.0%, transdermal agent water-solubleazone 2.0%, solvent dehydrated alcohol 79.0%.Preparation method is with embodiment 3.

Embodiment 7

Killing aquatic animal external parasite preventing and treating agent, mass percent is: Palmic acid 16.0%, surface active agent tween-205.0%, transdermal agent water-solubleazone 1.0%, solvent dehydrated alcohol 78.0%.Preparation method is with embodiment 3.

Embodiment 8

Killing aquatic animal external parasite preventing and treating agent, mass percent is: Palmic acid 20.0%, surface active agent tween-205.0%, transdermal agent water-solubleazone 4.0%, solvent dehydrated alcohol 71.0%.Preparation method is with embodiment 3.

Embodiment 9

Killing aquatic animal external parasite preventing and treating agent, mass percent is: Palmic acid 16.0%, surface active agent tween-207.0%, transdermal agent water-solubleazone 2.0%, solvent dehydrated alcohol 75.0%.Preparation method is with embodiment 3.

Embodiment 10

Killing aquatic animal external parasite preventing and treating agent, mass percent is: Palmic acid 1.0%, surface active agent tween-2010.0%, transdermal agent water-solubleazone 4.0%, solvent dehydrated alcohol 85.0%.Preparation method is with embodiment 3.

The pharmacodynamics test of embodiment 11 killing aquatic animal external parasite preventing and treating agent of the present invention

1 materials and methods

1.1 material

1.1.1 experimental animal

Host's model is Carassius auratus (Carassiusauratus), and parasite model is bad gill dactylogyrider (Dactylogyrusvastator); Infect Carassius auratus body weight 5.0 ± 0.5g.

1.1.2 for reagent product

Containing 16.0% Palmic acid external parasite preventing and treating agent for aquatic product animals, prepare by embodiment 9 formula.

1.1.3 instrument and equipment

XH-B vortex mixer: healthy medical apparatus company limited of Jiangyan City; BX41 optical microscope: OLYMPUS, Japan; Satorius ten thousand/electronic balance; Aquarium and temperature regulating device.

1.2 test method

Insecticidal test method as previously mentioned, is specially: 8 glass aquariums that use volume is 50cm × 40cm × 30cm, contains into abundant aeration tap water, pH7.0 ~ 7.5, controls water temperature 25 ± 1 DEG C, throws in Carassius auratus 100 tail infecting and have dactylogyrider respectively.First set control agent concentration on a large scale and carry out trial test, reset according to gradient increasing or decreasing according to parasite killing and the poisoning concentration of Carassius auratus again, add each acute drug respectively and stir, the film-making of depletion fish holobranch, microscopy during 48h, observe various sample killing effect to dactylogyrider under variable concentrations, statistics optimum effective concentration, the highest killing rate.Test repetition 3 times, and establish matched group.

2 results

2.1 matched group

The matched group fish that test is set up in carrying out is generally 100 tails, inspects 30 tails immediately by random samples respectively at 0h, 48h of test, dissects holobranch microscopy statistics and infects dactylogyrider par.At the 0h microscopy of test, dactylogyrider par is 56.2/tail, and 48h is carried out in test, and dactylogyrider par is 53.6/tail.Parasitic killing rate is calculated with the result of this and drug monitoring.

2.2 killing aquatic animal external parasite preventing and treating agents of the present invention kill result to dactylogyrider

16.0% Palmic acid killing aquatic animal external parasite preventing and treating agent, that kills dactylogyrider at 48h the results are shown in Table 5.As can be seen from Table 5, when concentration is 1.0mL/m ³time, insecticidal power is more weak, is only 61.2%; When concentration is 12.0mL/m ³time, its killing rate can reach 96.5%, and concentration is 15.0mL/m ³time killing rate be 100%; When concentration is 21.0mL/m ³time, still not there is intoxicating phenomenon in test fish, shows that said preparation is the good Biocidal fisheries drug of a kind of high-efficiency low-toxicity, application prospect.

Table 516.0% Palmic acid at 48h to dactylogyrider killing effect

Above-described embodiment is the present invention's preferably embodiment; but embodiments of the present invention are not restricted to the described embodiments; change, the modification done under other any does not deviate from spirit of the present invention and principle, substitute, combine, simplify; all should be the substitute mode of equivalence, be included within protection scope of the present invention.

Claims (9)

1. Palmic acid is preparing the application in killing aquatic animal vermin medicine.

2. application according to claim 1, is characterized in that: described aquatic animal is Fish.

3. application according to claim 1 and 2, is characterized in that: described parasite is dactylogyrider.

4. a killing aquatic animal external parasite preventing and treating agent, it is characterized in that being made up of the raw material of following masses percentage ratio: Palmic acid 1% ~ 20%, surfactant 5% ~ 10%, transdermal agent 1% ~ 4%, all the other are solvent, and the summation of above-mentioned raw materials is 100%.

5. killing aquatic animal external parasite preventing and treating agent according to claim 4, is characterized in that:

Described surfactant is tween 20.

6. killing aquatic animal external parasite preventing and treating agent according to claim 4, is characterized in that:

Described transdermal agent is water-solubleazone.

7. killing aquatic animal external parasite preventing and treating agent according to claim 4, is characterized in that:

Described solvent is dehydrated alcohol.

8. the preparation method of the killing aquatic animal external parasite preventing and treating agent described in any one of claim 4 ~ 7, is characterized in that comprising the following steps:

(1) Palmic acid, surfactant, transdermal agent, each raw material of solvent is taken, for subsequent use;

(2) Palmic acid and solvent are mixed to dissolve completely, obtain mixed solution;

(3) surfactant, transdermal agent are added in the mixed solution of step (2) gained stir, obtain killing aquatic animal external parasite preventing and treating agent.

9. the using method of the killing aquatic animal external parasite preventing and treating agent described in any one of claim 4 ~ 7, is characterized in that: the using dosage of described killing aquatic animal external parasite preventing and treating agent is 12 ~ 20mL/m ³.