CN108939614A - Polystyrene-diethylbenzene is without heating solid phase saponifiable extraction column and its preparation method and application - Google Patents
Polystyrene-diethylbenzene is without heating solid phase saponifiable extraction column and its preparation method and application Download PDFInfo
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- CN108939614A CN108939614A CN201810772213.4A CN201810772213A CN108939614A CN 108939614 A CN108939614 A CN 108939614A CN 201810772213 A CN201810772213 A CN 201810772213A CN 108939614 A CN108939614 A CN 108939614A
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- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01D—SEPARATION
- B01D15/00—Separating processes involving the treatment of liquids with solid sorbents; Apparatus therefor
- B01D15/08—Selective adsorption, e.g. chromatography
- B01D15/10—Selective adsorption, e.g. chromatography characterised by constructional or operational features
- B01D15/22—Selective adsorption, e.g. chromatography characterised by constructional or operational features relating to the construction of the column
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- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01D—SEPARATION
- B01D15/00—Separating processes involving the treatment of liquids with solid sorbents; Apparatus therefor
- B01D15/08—Selective adsorption, e.g. chromatography
- B01D15/10—Selective adsorption, e.g. chromatography characterised by constructional or operational features
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- B—PERFORMING OPERATIONS; TRANSPORTING
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- B01D15/08—Selective adsorption, e.g. chromatography
- B01D15/42—Selective adsorption, e.g. chromatography characterised by the development mode, e.g. by displacement or by elution
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- B01D15/426—Specific type of solvent
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- G—PHYSICS
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- 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
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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
- G01N30/04—Preparation or injection of sample to be analysed
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- G01N2030/062—Preparation extracting sample from raw material
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Abstract
The invention discloses a kind of polystyrene-divinylbenzenes without heating solid phase saponifiable extraction column and its preparation method and application, polystyrene-divinylbenzene includes void column pipe without heating solid phase saponifiable extraction column, exit sieve plate, polystyrene-divinylbenzene granular filler layer, isolation sieve plate and KOH powder filler layer are disposed in void column pipe from bottom to top, and the extraction column that the present invention is prepared can be applied to the sample pre-treatments containing lipid liquid.Compared with prior art, it is simple that the beneficial effects of the invention are as follows preparation methods, and when application significantly reduces saponification time, without additional heating and Solid Phase Extraction step, saves a large amount of time, improves the sensitivity of detection, improves the working efficiency of testing staff.
Description
Technical field
The invention belongs to extraction column preparation technical field, in particular to a kind of polystyrene-divinylbenzene is solid without heating
Phase saponifiable extraction column and its preparation method and application.
Background technique
It is saponified the Sample Pretreatment Technique Used as a kind of manifold, is usually used in releasing target analytes from lipid samples matrix
It releases, such as converts glycerol and fatty acid salt for glyceride, to reduce the load for entering organic phase in liquid-liquid extraction
Or interference.Saponification has been widely used in the sample treatment measured during some lyophobic dusts, and such as tocopherols are polycyclic
Arene, liposoluble vitamin class, phytosterol, carotenoid, mineral oil, Polychlorinated biphenyls, cholesterol etc..
Traditional liquid-liquid saponification usual reaction time is long, and heating water bath is needed to flow back or need using special instrument
Device is heated.The reaction temperature of traditional liquid-liquid saponification should generally be maintained at room temperature to this model of the boiling point of mixture
In enclosing, the reaction time is also from 30min to differing for 24 hours, or needs the longer reaction time.So traditional method for saponification both took
When it is laborious, and consumption of materials is big, it is more difficult to realize the fast and effective analysis to sample.
Summary of the invention
It is an object of the present invention to provide a kind of polyphenyl second for the above problem present in existing liquid-liquid saponification
Alkene-divinylbenzene is without heating solid phase saponifiable extraction column and its preparation method and application.
To achieve the goals above, the technical solution that the application uses are as follows:
A kind of polystyrene-diethylbenzene is without heating solid phase saponifiable extraction column, including void column pipe, in the void column pipe under
It is disposed with exit sieve plate, polystyrene-divinylbenzene granular filler layer, isolation sieve plate and KOH powder filler layer upwards,
The internal diameter of the void column pipe is 0.3cm~5cm, the height of the polystyrene-divinylbenzene granular filler layer be 0.5cm~
3cm, the height of the KOH powder filler layer are 0.5cm~3cm, the exit sieve plate and isolation sieve plate be absorbent cotton, filter paper,
One of mineral wool or porous ceramics.
Further, the void column pipe is plastic injector for temporary use or commodity-type solid-phase extraction column, and the void column pipe
Material be one of polypropylene, polyethylene or polytetrafluoroethylene (PTFE).
Further, the partial size of the polystyrene-divinylbenzene particle is 3 μm~100 μm, the polystyrene-
Divinylbenzene particles are that aperture isPorous particle or non-porous particle.
Preparation method of the polystyrene-diethylbenzene without heating solid phase saponifiable extraction column, includes the following steps:
S1, it takes a lower end to be equipped with the void column pipe of outlet, and is pressed into exit sieve plate to the bottom end of void column pipe；
S2, it filled polystyrene-divinylbenzene particles packing layer and is compacted in exit sieve plate；
S3, the indentation isolation sieve plate on polystyrene-divinylbenzene granular filler layer；
S4, it fills and KOH powder filler layer and is compacted on isolation sieve plate, the polystyrene-diethylbenzene is made without adding
Thermosetting phase saponifiable extraction column, and vacuum saves backup.
A kind of polystyrene-diethylbenzene is without heating solid phase saponifiable extraction column in the sample pre-treatments containing lipid liquid
Using.
Further, the polystyrene-diethylbenzene is used for the sample containing lipid liquid without heating solid phase saponifiable extraction column
The process of product pre-treatment is:
A, the sample containing lipid liquid is clicked and entered in the KOH powder filler layer without heating solid phase saponifiable extraction column, and be added dropwise
High purity water infiltrates extraction column, reacts 4min~10min；
B, extraction column is rinsed with high purity water again, after the liquid shows neutral flowed out, is eluted and extracted with 5mL~20mL organic solvent
It takes column 3~5 times, collects eluent, contain target analytes in the eluent；
C, after being evaporated the solvent in eluent, the organic solvent of 0.5mL~1.5mL is added, concussion dissolution crosses 0.22
μm filter membrane, be made analytical solution, for use.
Further, the sample containing lipid liquid be liquid fat, animal milk, blood, serum, dissolved with cream
Solution or solution dissolved with milk powder；The target analytes are cholesterol, phytosterol, liposoluble vitamin, fertility
Phenol, polycyclic aromatic hydrocarbon, carotenoid, Polychlorinated biphenyls or lycopene.
Further, the organic solvent is one or both of n-hexane, methanol, acetonitrile or isopropanol.
Compared with prior art, the beneficial effects of the present invention are:
(1) this polystyrene-diethylbenzene is not necessarily to heat when in use without heating solid phase saponifiable extraction column, user
Just.
(2) preparation method of this extraction column is simple, quick, safety and spends low.
(3) this extraction column has abandoned Traditional liquid phase in the application and has been saponified time-consuming in sample pretreatment process, laborious, expense
The disadvantages of solvent, significantly reduces saponification time, and is not necessarily to additional heating and Solid Phase Extraction (SPE) step, saves a large amount of
Time and hand labour, and can get good clean-up effect.
Detailed description of the invention
Fig. 1 is cholesterol standard curve (a) in the specific embodiment of the invention, the HPLC chromatogram of pig blood (b) and gutter oil (c)
Figure.
Specific embodiment
Technological means of the invention, creation characteristic, achieving the goal is easy to understand with effect in order to make, below in conjunction with
Technical solution of the present invention is clearly and completely described in the embodiment of the present invention.
Instrument and reagent used in the embodiment of the present invention are as follows:
Instrument: 1260 high performance liquid chromatograph of Agilent；MTN-2800D nitrogen evaporator；ART Miccra D-8 homogenizer；
MS3 basic model vortex mixer；BT 125D electronic balance.Reagent: acetonitrile (chromatographically pure), KOH, isopropanol, n-hexane (analysis
It is pure)；The ultrapure water prepared by Milli-Q ultrapure water instrument.
Standard substance: the cholesterol bought from Dr.Ehrenstorfer company.
Liquid phase chromatogram condition are as follows:
Chromatographic column: Agilent C18,250mm × 4.6mm, 5 μm；Mobile phase: acetonitrile/isopropanol (60/40, v/v)；Column
35 DEG C of temperature；Flow velocity 1.0mLmin-1；Detection wavelength 210nm；10 μ L of sample volume.
Embodiment 1
A kind of polystyrene-diethylbenzene is without heating solid phase saponifiable extraction column, including void column pipe, in void column pipe from bottom to top
It is disposed with exit sieve plate, polystyrene-divinylbenzene granular filler layer, isolation sieve plate and KOH powder filler layer, void column
The internal diameter of pipe is 0.3cm, and void column pipe is that material is polyacrylic plastic injector for temporary use, polystyrene-divinylbenzene
The height of particulate filler layer is 0.5cm, and polystyrene-divinylbenzene particle is the non-porous particle that partial size is 3 μm~100 μm, KOH
The height of powder filler layer is 0.5cm, and exit sieve plate and isolation sieve plate are absorbent cotton.
Above-mentioned polystyrene-diethylbenzene is as follows without heating solid phase saponifiable extraction column preparation process:
S1, it takes a lower end to be equipped with the void column pipe of outlet, and is pressed into exit sieve plate to the bottom end of void column pipe；
S2, it filled polystyrene-divinylbenzene particles packing layer and is compacted in exit sieve plate；
S3, the indentation isolation sieve plate on polystyrene-divinylbenzene granular filler layer；
S4, it fills and KOH powder filler layer and is compacted on isolation sieve plate, the polystyrene-diethylbenzene is made without adding
Thermosetting phase saponifiable extraction column, and vacuum saves backup.
Embodiment 2
A kind of polystyrene-diethylbenzene is without heating solid phase saponifiable extraction column, including void column pipe, in void column pipe from bottom to top
It is disposed with exit sieve plate, polystyrene-divinylbenzene granular filler layer, isolation sieve plate and KOH powder filler layer, void column
The internal diameter of pipe is 2.5cm, and void column pipe is the commodity-type solid-phase extraction column that material is polytetrafluoroethylene (PTFE), polystyrene-divinyl base
The height of benzene granular filler layer is 2.5cm, and polystyrene-divinylbenzene particle is non-porous that partial size is 3 μm~100 μm
Grain, the height of KOH powder filler layer are 2.5cm, and exit sieve plate and isolation sieve plate are mineral wool.
Above-mentioned polystyrene-diethylbenzene is same as Example 1 without heating solid phase saponifiable extraction column preparation process.
Embodiment 3
A kind of polystyrene-diethylbenzene is without heating solid phase saponifiable extraction column, including void column pipe, in void column pipe from bottom to top
It is disposed with exit sieve plate, polystyrene-divinylbenzene granular filler layer, isolation sieve plate and KOH powder filler layer, void column
The internal diameter of pipe is 5cm, and void column pipe is the commodity-type solid-phase extraction column that material is polyethylene, polystyrene-divinylbenzene particle
The height of packing layer is 3cm, and styrene-divinylbenzene particle is that partial size is 3 μm~100 μm porous particles, and aperture isThe height of KOH powder filler layer is 3cm, and exit sieve plate and isolation sieve plate are porous ceramics.
Above-mentioned polystyrene-diethylbenzene is same as Example 1 without heating solid phase saponifiable extraction column preparation process.
In order to further probe into the polystyrene-diethylbenzene prepared in above-described embodiment 1~3 without heating solid phase saponification
The application of extraction column, by taking polystyrene-diethylbenzene prepared by embodiment 2 is without heating solid phase saponifiable extraction column as an example, for containing
The sample pre-treatments of lipid liquid, process are as follows:
(1) preparation of sample
The 500 μ L of pig blood sample added with anti-coagulants is measured in test tube, and with normal saline dilution to 1.0mL, is made
Used time directly pipettes 100 μ L.
Trench oil samples do not need to handle, and when use directly pipettes 100 μ L.
(2) sample pre-treatments
A, sample is clicked and entered in the KOH powder filler layer of extraction column with liquid-transfering gun, and high purity water is added dropwise, soak extraction column
Profit, and react 6min；
B, extraction column is rinsed with high purity water again, after the liquid shows neutral flowed out, elutes extraction column 3 with 10mL n-hexane
It is secondary；
C, eluent is collected to reuse acetonitrile/isopropanol (60:40, v/v) after being dried with nitrogen solvent and be settled to 1mL,
Concussion dissolution, crosses 0.22 μm of filter membrane, analytical solution is made, for use.
The principle of self-heating solid phase saponification of the present invention is: water infiltration KOH powder filler layer is distilled when being added dropwise, solid KOH
Heat of solution can make barrel temperatures rise very rapidly up to 60~80 DEG C, while OH- concentration can be improved in excessive KOH, can promote soap
Change rapid reaction to carry out and (reacting completely in 5 minutes), while the KOH of high concentration can quickly promote the cell rupture in blood,
Quick release target analytes.
(3) preparation of standard working solution
Cholesterol standard substance 1mg is accurately weighed, is settled to 10mL with dehydrated alcohol, obtains 0.1mg/mL standard reserving solution, 4
The storage of DEG C dark place is stand-by.
Above-mentioned standard stock solution accurately is pipetted, is diluted step by step, 5 μ g/mL, 20 μ g/mL, 50 μ g/mL, 100 μ g/ are configured to
The cholesterol standard solution of mL, 200 μ g/mL and 500 μ g/mL.The standard working solution of various concentration is subjected to reversed-phase liquid chromatography
Method (RPLC) measurement carries out regression analysis to its respective concentration with the chromatographic peak area of standard working solution, and it is bent to obtain standard work
Line, such as table 1.
The standard curve of 1 cholesterol of table
(4) detection of sample and method performance evaluation
A, in sample cholesterol level detection
The sample solution Jing Guo preceding processing is being subjected to RPLC measurement under the same terms of measurement cholesterol standard curve,
The chromatographic peak area of cholesterol in blood sample is measured, standard curve is substituted into, obtains cholesterol level in sample liquid, then basis
The Mass Calculation of sample representated by sample solution obtains cholesterol residual quantity in sample.If cholesterol residual quantity is super in upper machine solution
The range of linearity upper limit is crossed, upper machine solution need to be diluted within the range of linearity with Extraction solvent.
B, method performance evaluation
A, recovery of standard addition and repeatability:
A certain amount of cholesterol standard solution is added in different blood samples (being shown in Table 2), above-mentioned blood is pressed after mixing
After sample pre-treatments step is handled, high performance liquid chromatography measurement is carried out.Measurement concentration and theory addition concentration are compared
Compared with obtaining sample TIANZHU XINGNAO Capsul, each pitch-based sphere is measured in parallel 6 times, obtains its relative standard deviation, measurement result is shown in Table 2.
As can be seen from Table 2, the average recovery rate of cholesterol is 97.6%~101.3%, and average relative standard's deviation (RSD) is
2.5%~3.7%, illustrate that the rate of recovery of the method for the present invention is higher, it is reproducible.
The recovery of standard addition and relative standard deviation RSD% of 2 cholesterol of table
B, detection limit:
The cholesterol matrix standard working solution of various concentration is injected into HPLC, with minimum concentration extraction standard solution chromatography
3 times of signal-to-noise ratio computation detection limits at peak, the detection of cholesterol are limited to 2 μ g/kg.
Fig. 1 gives above-mentioned 2 kinds of fluid samples containing cholesterol by this method treated high-efficient liquid phase chromatogram, wherein gallbladder
Completely and baseline separation, and chromatogram very " clean " shows this without heating self-heating saponification solid phase sterol target peak without miscellaneous peak
Pretreatment process of the extraction column for above two sample is successful.
In summary:
(1) this polystyrene-diethylbenzene is not necessarily to heat when in use without heating solid phase saponifiable extraction column, user
Just.
(2) preparation method of this extraction column is simple, quick, safety and spends low.
(3) this extraction column has abandoned Traditional liquid phase in the application and has been saponified time-consuming in sample pretreatment process, laborious, expense
The disadvantages of solvent, significantly reduces saponification time, and is not necessarily to additional heating and Solid Phase Extraction (SPE) step, saves a large amount of
Time and hand labour, and can get good clean-up effect.
Disclosed above is only presently preferred embodiments of the present invention, and still, the embodiment of the present invention is not limited to this, Ren Heben
What the technical staff in field can think variation should all fall into protection scope of the present invention.
Claims (8)
1. a kind of polystyrene-diethylbenzene is without heating solid phase saponifiable extraction column, which is characterized in that including void column pipe, the sky
Exit sieve plate, polystyrene-divinylbenzene granular filler layer, isolation sieve plate and KOH are disposed in column tube from bottom to top
Powder filler layer, the internal diameter of the void column pipe are 0.3cm~5cm, the height of the polystyrene-divinylbenzene granular filler layer
Degree is 0.5cm~3cm, and the height of the KOH powder filler layer is 0.5cm~3cm, and the exit sieve plate and isolation sieve plate are de-
One of rouge cotton, filter paper, mineral wool or porous ceramics.
2. polystyrene-diethylbenzene as described in claim 1 is without heating solid phase saponifiable extraction column, which is characterized in that described
Void column pipe is plastic injector for temporary use or commodity-type solid-phase extraction column, and the material of the void column pipe is polypropylene, polyethylene
Or one of polytetrafluoroethylene (PTFE).
3. polystyrene-diethylbenzene as described in claim 1 is without heating solid phase saponifiable extraction column, which is characterized in that described
The partial size of polystyrene-divinylbenzene particle is 3 μm~100 μm, and the polystyrene-divinylbenzene particle is that aperture isPorous particle or non-porous particle.
4. preparation method of the polystyrene-diethylbenzene as described in claim 1 without heating solid phase saponifiable extraction column, feature
It is, includes the following steps:
S1, it takes a lower end to be equipped with the void column pipe of outlet, and is pressed into exit sieve plate to the bottom end of void column pipe；
S2, it filled polystyrene-divinylbenzene particles packing layer and is compacted in exit sieve plate；
S3, the indentation isolation sieve plate on polystyrene-divinylbenzene granular filler layer；
S4, KOH powder filler layer is filled on isolation sieve plate and is compacted, it is solid without heating that the polystyrene-diethylbenzene is made
Phase saponifiable extraction column, and vacuum saves backup.
5. polystyrene-diethylbenzene as described in claim 1 is without heating solid phase saponifiable extraction column in the sample containing lipid liquid
Application in product pre-treatment.
6. application as claimed in claim 5, which is characterized in that the polystyrene-diethylbenzene is without heating solid phase saponification extraction
Taking process of the column for the sample pre-treatments containing lipid liquid is:
A, the sample containing lipid liquid is clicked and entered in the KOH powder filler layer without heating solid phase saponifiable extraction column, and be added dropwise high-purity
Water infiltrates extraction column, reacts 4min~10min；
B, extraction column is rinsed with high purity water again, after the liquid shows neutral flowed out, elutes extraction column with 5mL~20mL organic solvent
3~5 times, eluent is collected, contains target analytes in the eluent；
C, after being evaporated the solvent in eluent, the organic solvent of 0.5mL~1.5mL is added, concussion dissolution crosses 0.22 μm
Analytical solution is made, for use in filter membrane.
7. application as claimed in claim 6, which is characterized in that the sample containing lipid liquid be liquid fat, animal milk,
Blood, serum, the solution dissolved with cream or the solution dissolved with milk powder；The target analytes are cholesterol, plant steroid
Alcohols, liposoluble vitamin, tocopherol, polycyclic aromatic hydrocarbon, carotenoid, Polychlorinated biphenyls or lycopene.
8. application as claimed in claim 6, which is characterized in that the organic solvent is n-hexane, methanol, acetonitrile or isopropanol
One or both of.
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| CN110596270A (en) * | 2019-09-25 | 2019-12-20 | 浙江省疾病预防控制中心 | Solid phase extraction column for simultaneously detecting 8 tocopherol forms of vitamin A and vitamin E in food |
| US11447716B2 (en) * | 2019-05-23 | 2022-09-20 | Mark Herwig | Apparatus and method for acquiring essential oils |
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Cited By (2)
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| US11447716B2 (en) * | 2019-05-23 | 2022-09-20 | Mark Herwig | Apparatus and method for acquiring essential oils |
| CN110596270A (en) * | 2019-09-25 | 2019-12-20 | 浙江省疾病预防控制中心 | Solid phase extraction column for simultaneously detecting 8 tocopherol forms of vitamin A and vitamin E in food |
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Application publication date: 20181207 |