CN101949907A - Steroid pretreatment and detection method - Google Patents
Steroid pretreatment and detection method Download PDFInfo
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- CN101949907A CN101949907A CN2010102131786A CN201010213178A CN101949907A CN 101949907 A CN101949907 A CN 101949907A CN 2010102131786 A CN2010102131786 A CN 2010102131786A CN 201010213178 A CN201010213178 A CN 201010213178A CN 101949907 A CN101949907 A CN 101949907A
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Abstract
The invention relates to a steroid pretreatment and detection method. The method comprises the following steps: (1) sampling; (2) mixing a sample with extractant evenly to treat; (3) mixing the obtained supernatant and distilled water; (4) placing the mixed liquid in a special solid phase extraction column to perform solid phase extraction; (5) eluting the solid phase extraction column with abluent; (6) eluting the solid phase extraction column with eluant to obtain components under test; (7) obtaining eluent to treat with the microporous filter membrane; and (8) analyzing and determining the steroid components. By adopting the method of the invention, eleven steroid substances such as testosterone, methyltestosterone and methandrostenolone can be detected at once, the problem that multi-residue analysis can not be performed is solved; the novel extractant and special steroid solid phase extraction column are adopted, the extraction efficiency is high, the purification effect is good, the problems of low extraction efficiency and poor purification effect are solved; and the detection speed and accuracy are high, and better technical effect can be obtained.
Description
Technical field
The present invention relates to a kind of pre-treating method of steroids, belong to the food security field.
Background technology
The effect that promotes production of steroid medicine is many-sided, mainly show as the promotion protein synthesis, improve basic metabolism, improve feed conversion rate, reduce material consumption etc., it can strengthen substance in vivo deposition and improve production performance, can the remarkable and direct economic benefit of very fast generation, so the producer is had very big attractive force.Use hormone (non-therapeutical uses) to have nearly 50 years history in the animal husbandry, production of fodder producer, feed dealer and professional raising farmer that considerable part is arranged, be applied in the poultry farming as diethylstilbestrol, in premix and feed addictive, mix feed poultry, fowl or imbed the poultry basal part of the ear and the poultry wing under, to promote growth of animal and to improve livestock and poultry lean meat ratio.
If use more steroid drug main male sex hormone, estrogen, progestational hormone and many materials with estrogen effect in herding, breed industry.The more important steroid drug main of wherein residual meaning will comprise: vigorous earthworm (Boldenone, BOL), dehydroisoandrosterone (Dehydroepiandrosterone), methyltestosterone (Methyl testosterone, MTS), testobolin (Nandrolone propionate), the vigorous dragon of alcohol (Trenbolone, TRE), testosterone (Testosterone, 17 β TS), testosterone propionate (Testosterone propionate, PTS), stanozolol (Stanozlol), nandrolone (Nandrolone), progesterone (Progesterone), metandienone (Methandienone) and many material such as diethylstilbestrol (Diethylstilbestrol with estrogen effect, DES), hexestrol (Hexestrol, HES), dienestrol (Dienestrol, DIS), ZER (Zeranol, ZER), (Zearalenone ZLE) waits material to zearalenone.
Present detection method mainly contains methods such as LC-MS, gas chromatography mass spectrometry and immunoassay.Its pre-treatment process need through repeatedly extract, operations such as centrifugal, liquid-liquid extraction and rotary evaporation, process is loaded down with trivial details, be difficult for grasping, to the experimenter operate require high; The part test item need carry out enzymolysis, needs mostly to surpass 15 hours enzymolysis process, makes detection time very long; For miscellaneous steroid material, most of method can only detect at unitem, can not realize multi-residue analysis, and the detection cost is increased greatly; There are problems such as extraction efficiency is low, clean-up effect difference in addition.
Summary of the invention
A kind of pre-treating method of steroids preferably is provided, and its technical scheme is as following description:
A kind of steroid pre-treatment detection method comprises: the sampling of (1) sample; (2) sample and extraction agent are mixed and handle; (3) obtaining supernatant mixes with distilled water; (4) place mixed liquor and in special-purpose solid-phase extraction column, carry out Solid-Phase Extraction; (5) with washing agent drip washing solid-phase extraction column; (6) obtain component to be measured with eluant, eluent from solid-phase extraction column; (7) obtain eluent and handle through miillpore filter; (8) component of analysis and mensuration steroids.
Further, preferable methods is, during the sample sampling: solid sample is removed the peel, boned, smash sampling to pieces after the deburring; Semi-solid, the thickness sample is smashed to pieces or stir the back sampling; Fluid sample mixes the back direct sample.
Further, preferable methods is, in the step (2), the extraction agent supernatant be at first oscillation treatment, carry out centrifugal treating again after the ultrasonic Extraction then, obtain supernatant at last.
Further, preferable methods is to carry out Solid-Phase Extraction with 2～3mL/min flow velocity.
Further, preferable methods is that described detection method is that UPLC-MS-MS measures.
Further, described extraction agent is the potpourri of 99% acetonitrile and 1% acetic acid.
Further, described washing agent is the potpourri of 75% acetonitrile and 25% water.
Further, described washing agent is the potpourri of 50% methyl alcohol and 50% water.
Further, described solid-phase extraction column is the C18 post.
This method reduced complicated centrifugal, revolve the number of times of operations such as steaming, simplified operating process, shortened in extraction time to one hour, solved long problem of pre-treatment time; The step that simplifies the operation is as long as the operation of seven steps just can be finished the pre-treatment process, loaded down with trivial details, the not wield problem of the process that solved; Can one-time detection steroid material in the vigorous dragon of alcohol, methyltestosterone, metandienone etc. 11, solved can not multi-residue analysis problem; Pre-treatment detection method of the present invention, the extraction efficiency height, good purification has solved that extraction efficiency is low, the problem of clean-up effect difference; The fast precision height of detection speed has better technical effect.
Description of drawings
Below in conjunction with accompanying drawing the present invention is carried out further detailed description, so that above-mentioned advantage of the present invention is clearer and more definite.
Fig. 1 is the process flow diagram of steroids pre-treatment detection method of the present invention.
Embodiment
The present invention is described in further detail below in conjunction with accompanying drawing.
As shown in the figure, the present invention handles according to following flow process: at first carry out sample sampling, (solid sample is removed the peel, is boned, smash sampling to pieces after the deburring accurately to take by weighing the sample 5.0g that handled well; Semi-solid, the thickness sample is smashed to pieces or stir the back sampling; Fluid sample mixes the back direct sample) in the 50mL centrifuge tube.
Next, above-mentioned sample is handled, extraction agent (liquid 20.0mL) in the adding steroid fast detecting pretreatment reagent kit vibrates in vortex mixer and extracts 1min, ultrasonic Extraction 20min in ultrasonic generator, with the centrifugal 5min of 12000r/min, take out the 5mL supernatant in centrifuge tube, add 10mL distilled water and mix.
Then, place mixed liquor in extraction column and Solid-Phase Extraction; With washing agent drip washing solid-phase extraction column.Get mixed liquor and in the special-purpose solid-phase extraction column of steroid, carry out Solid-Phase Extraction with 2～3mL/min flow velocity, treat that sample liquid is crossed post after, with 5mL washing agent drip washing solid-phase extraction column.
Obtain component to be measured with eluant, eluent from solid-phase extraction column; Obtain eluent and handle through miillpore filter.Discard leacheate, component to be measured is eluted from solid-phase extraction column, receive whole eluents, mix, cross 0.20 μ m miillpore filter with the 2mL eluant, eluent.
At last, utilize UPLC-MS-MS analysis of test methods and measure the component of steroids.
Wherein, described extraction agent is the potpourri of 99% acetonitrile and 1% acetic acid; Described washing agent is the potpourri of 75% acetonitrile and 25% water; Described washing agent is the potpourri of 50% methyl alcohol and 50% water; Described solid-phase extraction column is the extraction column of C18 post or other similar functions.
This method reduced complicated centrifugal, revolve the number of times of operations such as steaming, simplified operating process, shortened in extraction time to one hour, solved long problem of pre-treatment time; The step that simplifies the operation is as long as the operation of seven steps just can be finished the pre-treatment process, loaded down with trivial details, the not wield problem of the process that solved; Can one-time detection steroid material in the vigorous dragon of alcohol, methyltestosterone, metandienone etc. 11, solved can not multi-residue analysis problem; Pre-treatment detection method of the present invention, the extraction efficiency height, good purification has solved that extraction efficiency is low, the problem of clean-up effect difference; The fast precision height of detection speed has better technical effect.
Described specific embodiment only is exemplary, and under above-mentioned instruction of the present invention, those skilled in the art can carry out various improvement and distortion on the basis of the foregoing description, and these improvement or distortion drop in protection scope of the present invention.It will be understood by those skilled in the art that top specific descriptions just in order to explain purpose of the present invention, are not to be used to limit the present invention.Protection scope of the present invention is limited by claim and equivalent thereof.
Claims (9)
1. a steroid pre-treatment detection method comprises: the sampling of (1) sample; (2) sample and extraction agent are mixed and handle; (3) obtaining supernatant mixes with distilled water; (4) place mixed liquor and in special-purpose solid-phase extraction column, carry out Solid-Phase Extraction; (5) with washing agent drip washing solid-phase extraction column; (6) obtain component to be measured with eluant, eluent from solid-phase extraction column; (7) obtain eluent and handle through miillpore filter; (8) component of analysis and mensuration steroids.
2. steroid pre-treatment detection method according to claim 1 is characterized in that, sample when sampling: solid sample is removed the peel, is boned, smash sampling to pieces after the deburring; Semi-solid, the thickness sample is smashed to pieces or stir the back sampling; Fluid sample mixes the back direct sample.
3. steroid pre-treatment detection method according to claim 1 is characterized in that, in the step (2), the extraction agent supernatant be at first oscillation treatment, carry out centrifugal treating again after the ultrasonic Extraction then, obtain supernatant at last.
4. steroid pre-treatment detection method according to claim 1 is characterized in that, carries out Solid-Phase Extraction with 2～3mL/min flow velocity.
5. steroid pre-treatment detection method according to claim 1 is characterized in that, described detection method is that UPLC-MS-MS measures.
6. steroid pre-treatment detection method according to claim 1 is characterized in that described extraction agent is the potpourri of 99% acetonitrile and 1% acetic acid.
7. steroid pre-treatment detection method according to claim 1 is characterized in that described washing agent is the potpourri of 75% acetonitrile and 25% water.
8. steroid pre-treatment detection method according to claim 1 is characterized in that described washing agent is the potpourri of 50% methyl alcohol and 50% water.
9. steroid pre-treatment detection method according to claim 1 is characterized in that described solid-phase extraction column is the C18 post.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2010102131786A CN101949907A (en) | 2010-06-30 | 2010-06-30 | Steroid pretreatment and detection method |
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| Application Number | Priority Date | Filing Date | Title |
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| CN2010102131786A CN101949907A (en) | 2010-06-30 | 2010-06-30 | Steroid pretreatment and detection method |
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| CN101949907A true CN101949907A (en) | 2011-01-19 |
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| CN2010102131786A Pending CN101949907A (en) | 2010-06-30 | 2010-06-30 | Steroid pretreatment and detection method |
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Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US3041245A (en) * | 1958-09-10 | 1962-06-26 | Thomae Gmbh Dr K | Composition for the care of the skin |
| CN1526721A (en) * | 2003-03-06 | 2004-09-08 | 涛 周 | Extraction process of conjugated female hormone |
-
2010
- 2010-06-30 CN CN2010102131786A patent/CN101949907A/en active Pending
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US3041245A (en) * | 1958-09-10 | 1962-06-26 | Thomae Gmbh Dr K | Composition for the care of the skin |
| CN1526721A (en) * | 2003-03-06 | 2004-09-08 | 涛 周 | Extraction process of conjugated female hormone |
Non-Patent Citations (7)
| Title |
|---|
| ANNE PIRAM ET.AL.: "《Development and optimisation of a single extraction procedure for the LC/MS/MS analysis of two pharmaceutical classes residues in sewage treatment plant》", 《TALANTA》 * |
| CAROLE KELLY: "《Analysis of steroids in environmental water samples using solid-phase extraction and ion-trap gas chromatography一mass spectrometry and gas chromatography一tandem mass spectrometry》", 《JOURNAL OF CHROMATOGRAPHY A》 * |
| MARIA J. LOPEZ DE ALDA ET.AL.: "《Determination of steroid sex hormones and related synthetic compounds considered as endocrine disrupters in water by fully automated on-line solid-phase extraction-liquid chromatography- diode array detection》", 《JOURNAL OF CLU-OMATOGRAPHY A》 * |
| 吕惠卿等: "《固相萃取-液相色谱串联质谱同时测定牛奶中8种甾类同化激素多残留》", 《食品科学（分析检测）》 * |
| 夏敏等: "《高效液相色谱串联质谱法测定动物源食品中群勃龙、勃地酮和黄体酮残留量》", 《分析测试学报》 * |
| 崔晓亮等: "《超高效液相色谱-串联电喷雾四级杆质谱法同时测定牛奶中12中糖皮质激素的残留》", 《色谱》 * |
| 徐锦忠等: "《高效液相色谱-串联质谱法同时检测鸡肉和鸡蛋中合成类固醇类激素和糖皮质激素》", 《分析化学研究报告》 * |
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Application publication date: 20110119 |