CN110031562A - Pi Lebao raw medicine is as the application for remaining marker in grass carp tissue - Google Patents
Pi Lebao raw medicine is as the application for remaining marker in grass carp tissue Download PDFInfo
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- CN110031562A CN110031562A CN201910344658.7A CN201910344658A CN110031562A CN 110031562 A CN110031562 A CN 110031562A CN 201910344658 A CN201910344658 A CN 201910344658A CN 110031562 A CN110031562 A CN 110031562A
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
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Abstract
The invention discloses Pi Lebao raw medicines as the application for remaining marker in grass carp tissue, through studying, Pi Lebao raw medicine is in liver, kidney, muscle and intestinal tissue, the release rate of Pi Lebao raw medicine is most slow, residual quantity proportion highest, therefore using Pi Lebao raw medicine as the residual marker in grass carp tissue.
Description
Technical field
The present invention relates to Pi Lebao raw medicines as the application for remaining marker in grass carp tissue.Belong to drug measurement techniques neck
Domain.
Background technique
Pi Lebao is being commonly called as bronopol, and full name is 2- bromo-2-nitro-1,3-propylene glycol, be under room temperature white extremely
Faint yellow, yellowish-brown crystalline powder, odorless, tasteless, soluble easily in water, ethyl alcohol, propylene glycol are insoluble in chloroform, acetone, benzene etc..
It will be slow decomposition in alkaline aqueous solution, have corrosiveness to certain metals, such as aluminium.Bronopol is toxic, to eyes, skin, glues
The irritating effect of film, can cause birth defect, a large amount of to use, to environment nocuousness.Main application is the preservative of cosmetics, is killed
Bacteriocin can effectively prevent various plants pathogenetic bacteria.
In aquaculture field, since bronopol has high-efficiency broad spectrum antibacterial action, it is commonly used for helminth and kills and be fertilized
Ovum disinfection, but in practical applications, nothing is related in aquaculture for having hazard residue in animal body, but both at home and abroad for bronopol
The standard of interior medicament residue limitation in animal body.In Japan, bronopol allows the disinfectant as clupeoids fertilized eggs to make
With in addition, Japanese relevant department treats the development experiments of salmon class saprolegniasis in development with bronopol.In US and European
Etc. ground, bronopol be chiefly used in water treatment agent, it is antibacterial and kill harmful bacteria in water.China Sun Peng et al. inhibits rainbow using bronopol
Trout fertilized eggs water mo(u)ld, the results showed that when bronopol concentration is higher than 300mg/ml, fungistatic effect is significant.Domestic Tao Jilai et al.,
Toxic reaction, toxicity target organ and the possible delay of animal after observation bronopol is given in a single dose and gives repeatedly SD rat 90 days
Toxicity, the results show that the LOAEL (minimum dose of side reaction occur) that bronopol solution orally contaminates 90 days daily to rat is
40mg/kg/day, NOAEL (dosage level for not occurring side reaction) are 20mg/kg/day, do not find target organ.Bronopol without
Obvious teratogenesis mutagenesis, parent and embryo-fetus development dosage that has no adverse reaction (NOAEL) to rat are 35mg/
kg/day.Therefore bronopol residue problem has to attract people's attention in aquatic biological.Grass carp is the four of CHINESE FRESHWATER cultivation
One of large Chinese carp, rapidly, feed resource is wide for growth.Grass carp is liable to illness, unavoidably will use Pi Lebao in the breeding process
Drug inside, if residual quantity is exceeded will necessarily to cause adverse effect to human health, therefore monitoring medicament residue is very must
It wants.It there is no Pi Lebao remaining correlative study in grass carp tissue at present, remain mark in tissue using Pi Lebao raw medicine as grass carp
Will object determines residue limits of the Pi Lebao in grass carp using high performance liquid chromatography, is Pi Lebao in grass carp breeding process
Operating specification provide foundation.
At present there is no Pi Lebao grass carp tissue in remaining correlative study, Pi Lebao raw medicine as grass carp tissue in remain
The application of marker is very important, and determines residue limits of the Pi Liaobao in grass carp using high performance liquid chromatography.
Summary of the invention
The purpose of the present invention is to overcome above-mentioned the deficiencies in the prior art, provide Pi Lebao raw medicine as residual in grass carp tissue
Stay the application of marker.
To achieve the above object, the present invention adopts the following technical solutions:
Pi Lebao raw medicine, i.e. 2- bromo-2-nitro-1,3-propylene glycol, as the application for remaining marker in grass carp tissue.
Preferably, the residual target tissue of Pi Lebao raw medicine is liver.
Beneficial effects of the present invention:
Through studying, Pi Lebao raw medicine is in liver, kidney, muscle and intestinal tissue, and the release rate of Pi Lebao raw medicine is most
Slowly, residual quantity proportion highest, therefore using Pi Lebao raw medicine as the residual marker in grass carp tissue.
Detailed description of the invention
Fig. 1 is 6 hours sample chromatogram figures of liver；
Fig. 2 is 7 days sample chromatogram figures of liver；
Fig. 3 is 6 hours sample chromatogram figures of kidney；
Fig. 4 is 3 days sample chromatogram figures of kidney；
Fig. 5 is 6 hours sample chromatogram figures of muscle；
Fig. 6 is 3 days sample chromatogram figures of muscle；
Fig. 7 is 6 hours sample chromatogram figures of enteron aisle；
Fig. 8 is 3 days sample chromatogram figures of enteron aisle.
Specific embodiment
The present invention will be further elaborated with reference to the accompanying drawings and examples, it should which explanation, following the description is only
It is not to be defined to its content to explain the present invention.
Embodiment:
Pi Lebao control substance of plant drug: be purchased from Beijing Century AudioCodes Bioisystech Co., Ltd, German Dr.Ehrenstorfer,
CAS number 52-51-7, water content 0.3%, purity 98.5%, tolerance/uncertainty +/- 1.0%, karr Brigit Fischer (Schmidt) titration
Measurement.
Grass carp mouthful fills happy precious to coating, and dosage 200mg/kg takes corresponding mouth liquid filling once to be gavaged with conduit.6h and 1,
3,7,14d takes 1 group of (10 tail) grass carp at random respectively, and slaughter takes liver, kidney, muscle and intestinal samples respectively.It is stored in -20
DEG C, it is spare.
Pi Lebao main metabolites 2- nitro-1,3-propylene glycol (Br 1), the intracorporal residual content of grass carp >=
95.0%, other metabolites can't be residual marker less than 5%, there is display (mark in following residual eliminating figure
Object: being residence time longest drug in target tissue).The above metabolite is synthesized by this upper laboratory, and synthetic method is as follows:
20mL methanol is added in 100mL there-necked flask, opens stirring, and metallic sodium 0.26g is added to 5 DEG C in ice salt bath temperature control
(11.3mmol) waits hydrogen release to finish, is slowly warming up to 40 DEG C and keeps the temperature 3 hours, be then cooled to 5 DEG C of addition paraformaldehydes
0.6g (20mmol) is then cooled to 0 DEG C of methanol solution 40ml for starting that nitromethane 0.61g (10mmol) is added dropwise, and is added dropwise
Control temperature is no more than 5 DEG C in the process, after being added dropwise 5 DEG C insulation reaction 24 hours, filtering, solid washed with cold methanol 10ml
It washs twice, (25-30 DEG C/10mmHg) of vacuum drying obtains intermediate 1.8g, be dissolved into 18mL methyl- tert in this intermediate 1.8g
In butyl ether, the lower solution that 1g salicylic acid is added dropwise and is dissolved into 10ml methyl tertiary butyl ether is stirred, is heated to 40-45 after stirring 4 hours
DEG C reaction 1 hour, be then cooled to room temperature, filter, filtrate recycling methyl tertiary butyl ether(MTBE) to 2mL then placement refrigerator be cooled to-
10 DEG C, solid filtering is obtained, drying obtains product 0.4g, the main metabolites 2- nitro-1,3-propylene glycol of as Pi Lebao.
Synthetic route is as follows:
Grass carp tissue 5g (being accurate to 0.01g) is accurately weighed, 17.5mL bronopol special extract (article No.: OR- is added
1801,500mL/ bottles, it is purchased from Ningbo Ou Pu Instrument Ltd.) and 10mL n-hexane, high-speed homogenization 5min, vortex 5min,
4500r/min is centrifuged 10min, takes middle layer liquid to cross miillpore filter, to upper machine.
Chromatographic condition: Agilent ZORBAX SB-C18 column (250mm × 4.6mm, 5 μm)；With phosphoric acid water (phosphoric acid/water=
1/1000, V/V): methanol=95:5 (V:V) is mobile phase；Flow velocity 1mL/min, Detection wavelength 210nm；30 DEG C of temperature of detection.Group
Drug concentration in tissue samples, which directly measures, to be obtained.Experimental data is handled and is analyzed by SPSS (16.0 version) software.
1, the residual distribution of Pi Lebao and its metabolite in grass carp is respectively organized:
Pi Lebao is distributed in Liver of Ctenopharyngodon Idellus, kidney, muscle and enteron aisle.6h~7d after administration, drug is dense in liver
Degree is above kidney, muscle and intestinal tissue, reaches as high as 3515.23 μ g/kg；To 14d, above four kinds of tissues be can't detect
Pi Lebao residual, is shown in Table 1.7d upon administration, Pi Lebao in the tissue residual quantity sequence be liver > kidney > enteron aisle > muscle.
Result above prompt, the organ that Pi Lebao finally leaves is liver, preliminary to infer that liver is its remaining target tissue.
Concentration of 1. Pi Lebao of table in grass carp is organized
ND: it lower than detection limit or is not detected.
2, the composition of metabolite in respectively organizing:
Metabolite in liver: 3d after drug withdrawal can detect Pi Lebao (Br0) and its metabolite in Liver of Ctenopharyngodon Idellus
(Br1).7d only detects original shape drug；(table 2) is not detected in 14d original shape medicine and metabolite.6h sample chromatogram figure is shown in
Fig. 1,7d sample chromatogram figure are shown in Fig. 2.
Pi Lebao and its metabolite in 2. Liver of Ctenopharyngodon Idellus of table
ND: it lower than detection limit or is not detected.
Metabolite in kidney: 1d after drug withdrawal can detect Pi Lebao (Br0) and its metabolite in grass carp kidney
(Br1).3d only detects original shape drug (Br0).(see Table 3 for details) is not detected in 14d original shape medicine and metabolite.6h sample
Product chromatogram is shown in that Fig. 3,3d sample chromatogram figure are shown in Fig. 4.
Pi Lebao and its metabolite in 3. grass carp kidney of table
ND: it lower than detection limit or is not detected.
Metabolite in muscle: 1d after drug withdrawal can detect Pi Lebao (Br0) and its metabolite in grass carp muscle
(Br1).3d only detects original shape drug (Br0).(see Table 4 for details) is not detected in 14d prototype medicine and metabolite.6h sample
Product chromatogram is shown in that Fig. 5,3d sample chromatogram figure are shown in Fig. 6.
Pi Lebao and its metabolite in 4 grass carp muscle of table
ND: it lower than detection limit or is not detected.
Metabolite in enteron aisle: 6h after drug withdrawal, grass carp intestinal can detect that Pi Lebao raw medicine (Br0) and its metabolism produce
Object (Br1).3d only detects original shape drug (Br0).(table 5) is not detected in 14d prototype medicine and metabolite.6h sample color
Spectrogram is shown in that Fig. 7,3d sample chromatogram figure are shown in Fig. 8.
Pi Lebao and its metabolite in 5. grass carp intestinal of table
ND: it lower than detection limit or is not detected.
In 6h, grass carp respectively organize in ratio of the Pi Lebao raw medicine (Br0) in liver, kidney and enteron aisle be above 90%,
The ratio of Pi Lebao raw medicine (Br0) is up to 97.37% in muscle.Detailed results are shown in Table 6.According to 1~table of table 6 as a result, Pi Lebao
And its metabolin residual quantity highest in liver, release rate is most slow, therefore liver is its residual target tissue.
The ratio (6h) of Pi Lebao and its metabolite and total residue in 6. grass carp of table tissue
The tissue of Pi Lebao and its metabolite is eliminated:
Pi Lebao and its metabolite main residual eliminating parameter in grass carp is organized are shown in Table 7.Pi Lebao raw medicine (Br0) exists
Half-life period in four kinds of tissues, the release rate in liver was most slow, and half-life period is between 54.24h~51.17h
54.24h also further confirms that liver is its residual target tissue.
In residual target tissue --- in liver, the half-life period of Pi Lebao metabolite (Br1) is 13.25h., is far below skin
The half-life period of happy treasured raw medicine (Br0).In kidney, muscle and intestinal tissue, the half-life period of Pi Lebao metabolite (Br1) is also remote
Lower than the half-life period of Pi Lebao raw medicine (Br0).Therefore, Pi Lebao raw medicine (Br0) is Pi Lebao in the intracorporal residual mark of grass carp
Object.
Main residual eliminating parameter in 7 different tissues of table
Pi Lebao is distributed in Liver of Ctenopharyngodon Idellus, kidney, muscle and intestinal tissue；The sequence of residual quantity is liver > kidney
Dirty > enteron aisle > muscle；Pi Lebao and its metabolin the residual quantity highest in liver, release rate is most slow, therefore liver is the residual of its
Stay target tissue.Either in residual target tissue liver, or in kidney, muscle and intestinal tissue, Pi Lebao raw medicine (Br0) disappears
Except speed is most slow, residual quantity proportion highest, the half-life period of Pi Lebao metabolite (Br1) and residual quantity ratio are far below
Pi Lebao raw medicine (Br0).Therefore, Pi Lebao raw medicine (Br0) is Pi Lebao in the intracorporal residual marker of grass carp.
Above-mentioned, although the foregoing specific embodiments of the present invention is described with reference to the accompanying drawings, not protects model to the present invention
The limitation enclosed, based on the technical solutions of the present invention, those skilled in the art are not needed to make the creative labor and can be done
Various modifications or changes out are still within protection scope of the present invention.
Claims (2)
1. Pi Lebao raw medicine is as the application for remaining marker in grass carp tissue.
2. application as described in claim 1, it is characterized in that: the residual target tissue of Pi Lebao raw medicine is liver.
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201910344658.7A CN110031562A (en) | 2019-04-26 | 2019-04-26 | Pi Lebao raw medicine is as the application for remaining marker in grass carp tissue |
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| CN201910344658.7A CN110031562A (en) | 2019-04-26 | 2019-04-26 | Pi Lebao raw medicine is as the application for remaining marker in grass carp tissue |
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Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US6160023A (en) * | 1996-08-01 | 2000-12-12 | Vericore Limited | Use of bronopol for the treatment of diseases in fish |
| CN104267125A (en) * | 2014-10-13 | 2015-01-07 | 中国水产科学研究院长江水产研究所 | Method for predicting drug residues in grass carp tissue with physiological pharmacokinetic model |
| CN106431962A (en) * | 2016-06-23 | 2017-02-22 | 上海海洋大学 | Metabolites of Ridomil in ctenopharyngodon idellus |
-
2019
- 2019-04-26 CN CN201910344658.7A patent/CN110031562A/en active Pending
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US6160023A (en) * | 1996-08-01 | 2000-12-12 | Vericore Limited | Use of bronopol for the treatment of diseases in fish |
| CN104267125A (en) * | 2014-10-13 | 2015-01-07 | 中国水产科学研究院长江水产研究所 | Method for predicting drug residues in grass carp tissue with physiological pharmacokinetic model |
| CN106431962A (en) * | 2016-06-23 | 2017-02-22 | 上海海洋大学 | Metabolites of Ridomil in ctenopharyngodon idellus |
Non-Patent Citations (5)
| Title |
|---|
| D.J.MORRIS 等: "EU sampling strategies for the detection of veterinary drug residues in aquaculture species: Are they working?", 《DRUG TESTING AND ANALYSIS》 * |
| DANIEL P.等: "Available chemotherapy in Mediterranean fish farming:use and needs", 《CIHEAM》 * |
| 梁赤周 等: "20%溴硝醇可湿性粉剂高效液相色谱分析方法研究", 《农药科学与管理》 * |
| 潘晓东 等: "超高效液相色谱-串联质谱法同时测定鱼肉中氯霉素、甲砜霉素和氟甲砜霉素", 《中国食品温室杂志》 * |
| 霍红 等: "《食品商品学》", 30 April 2015, 中国财富出版社 * |
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