CN103383321A - Fenton-method sample pretreatment method for metal-heteroatom zeolite molecular sieves - Google Patents
Fenton-method sample pretreatment method for metal-heteroatom zeolite molecular sieves Download PDFInfo
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Abstract
The invention discloses a Fenton-method sample pretreatment method for metal-heteroatom zeolite molecular sieves. The method comprises the steps of synthesizing metal-heteroatom ZSM-5 zeolite molecular sieves; adding the metal-heteroatom ZSM-5 zeolite molecular sieves into centrifugal tubes, and adding a biological sample solution and a H2O2 solution; transferring the centrifugal tubes into a thermostatic waterbath, inserting ultrasonic reactor probes to the certain depth below a liquid surface, and stopping after reacting for certain time; taking out the centrifugal tubes, centrifuging, transferring and taking supernate, and waiting to be tested; according to maximum absorbencies which are measured by an ultraviolet visible spectrophotometer before and after the protein reaction, combining a gel electrophoresis technology and Image-Lab software to perform quantitative analysis and measurement. By the aid of the method, complex actual samples, such as blood and protein solutions, are decomposed in advance in an efficient, fast, pollution-free and simple mode, so that requirements of fields such as subsequent instrumental analysis and testing can be met.
Description
Technical field
The invention belongs to technical field of function materials, relate to a kind of metal heteroatom zeolite molecular sieve Fenton method sample-pretreating method.
Background technology
Actual biological sample (as whole blood, dairy produce etc.) background constituent is complicated, and the organic content ratios such as protein are high, can not directly satisfy the analysis instrument test needs such as atomic spectrum, must carry out effective sample pretreatment.Traditional Wet Sample Pretreatment Technique consumes a large amount of severe corrosive acid, base reagent in process, may affect experiment operator health, does not meet modern times " green laboratory " theory.
Utilize Fenton reaction decomposes organism to have efficient high, reaction is at room temperature carried out, handling safety, the advantage such as be easy to get that agents useful for same is easy.The Fenton reaction is main generates hydroxyl radical free radical (OH) by the ferric ion catalyzing hydrogen peroxide, and OH further carries out oxidative dehydrogenation, addition reaction or change its cloud density and structure etc. organism making organic component be degraded to CO
2, H
2O。Recently, the people such as C.Phillip Shelor are to be entitled as " Fenton Digestion of Milk for Iodinalysis " (Fenton's reaction is cleared up in advance powdered milk sample and measured I), be published in " Analytical Chemistry " (U.S.'s analytical chemistry), in September, 2011, the 83rd volume, the page number: the 8300-8307 page), this research work utilizes ferric ion free in solution as the actual powdered milk sample of homogeneous phase Fenton catalysts resolution process.But it is worthy of note, exist free active iron ion easily to precipitate in the respective reaction medium, the not reproducible utilization of catalyzer is for harsh " bottleneck " problem that waits of the pH value requirement of example reaction medium.The heteroatomic ZSM-5 molecular sieve of Nano zeolite of doping metals in skeleton structure, micromechanism is enriched controlled, and it obtains extensive concern as effective heterogeneous Fenton reaction active catalyst.
In sum, the present invention is according to the deficiency of existing actual biological sample pretreatment technology, ultrasonic assistant metal heteroatoms zeolite molecular sieve Fenton method Sample Pretreatment Technique is proposed first, efficiently, quick, pollution-free, clear up in advance the complicated actual samples such as blood, protein solution easily, development provides the Sample Pretreatment Technique of " novel green safety ", to satisfy the field needs such as subsequent instrumentation analytical test.
Summary of the invention
For defective of the prior art, the purpose of this invention is to provide a kind of metal heteroatom zeolite molecular sieve Fenton method sample-pretreating method, degradation efficiency is high, and detection speed is fast, and the consumption amount of reagent is few, and is little to environmental hazard.
The present invention is achieved by the following technical solutions:
The invention provides a kind of metal heteroatom zeolite molecular sieve Fenton method sample-pretreating method, described method comprises the steps:
Step 1, metal heteroatom ZSM-5 zeolite molecular sieve synthetic;
Step 2 adds described metal heteroatom ZSM-5 zeolite molecular sieve in centrifuge tube, and adds wherein biological sample solution and H
2O
2Solution;
Step 3 is transferred to centrifuge tube in water bath with thermostatic control, the ultrasonic reactor probe is inserted under liquid level reaction;
Step 4 after question response is completed, is taken out centrifuge tube, and is centrifugal, pipettes supernatant, to be measured;
Step 5 records maximum absorbance before and after proteins react according to ultraviolet-visual spectrometer, utilizes sodium dodecyl sulfate-polyacrylamide gel electrophoresis technology and Image-Lab software to carry out quantitative test and mensuration to the band intensity before and after proteins react.
Preferably, in step 1, described metal heteroatom ZSM-5 zeolite molecular sieve is Fe-ZSM-5 zeolite molecular sieve or M-Fe-ZSM-5 zeolite molecular sieve, and wherein M is Cu, Mn or Ti.
Preferably, the synthetic of described Fe-ZSM-5 zeolite molecular sieve is specially: at first with Fe
2(SO
4)
39H
2The O stirring and dissolving is at H
2SO
4In solution, then slowly be added dropwise to the good Na of dissolving in advance under vigorous stirring
2SiO
39H
2O solution adds the tetrapropylene ammonium bromide that measures ratio after fully stirring, and stirs, and obtains aqueous precursor gel; The gained aqueous precursor gel is sealed under 180 ℃ reacts 96h, through cooling, filtration, after washing, drying, be transferred to roasting under 550 ℃ of conditions in muffle furnace.
Preferably, in described Fe-ZSM-5 zeolite molecular sieve, weight of iron number percent is 1～10%.
Preferably, the synthetic of described M-Fe-ZSM-5 zeolite molecular sieve is specially: at first with Fe
2(SO4)
39H
2O, sulfuric acid M salt are by measuring than stirring and dissolving at H
2SO
4In solution, then slowly be added dropwise to the good Na of dissolving in advance under vigorous stirring
2SiO
39H
2O solution adds the tetrapropylene ammonium bromide (TPABr) that measures ratio after fully stirring, and stirs, and obtains aqueous precursor gel; The gained aqueous precursor gel is sealed under 180 ℃ reacts 96h, through cooling, filtration, after washing, drying, be transferred to roasting under 550 ℃ of conditions in muffle furnace.
Preferably, in step 2, described biological sample solution is specially: with contained albumin, haemoglobin, fibrinogenic content ratio in blood, with the formulated finite concentration protein solution of NaCl solution dissolving, wherein the volumetric molar concentration of NaCl solution is 0.15moL/L respectively.
Preferably, in step 2, the pH value of described biological sample solution is 2.5～7.5, and it is 1moL/L HCl and 1moL/L NaOH that the pH of described biological sample solution regulates reagent.
Preferably, in step 2, described H
2O
2The solution quality percent concentration is 30%.
Preferably, in step 2, the mass volume ratio of described metal heteroatom ZSM-5 zeolite molecular sieve and described biological sample solution is (1～10): 1, and described mass volume ratio is: the metal heteroatom ZSM-5 zeolite molecular sieve that adds (1～10) mg in every 1mL biological sample solution; Described metal heteroatom ZSM-5 zeolite molecular sieve and described H
2O
2The mass volume ratio of solution is (100～1000): 1, and described mass volume ratio is: often add 1mL H
2O
2Solution is the metal heteroatom ZSM-5 zeolite molecular sieve of corresponding adding of (100～1000) mg.
Preferably, the mass volume ratio of described metal heteroatom ZSM-5 zeolite molecular sieve and described biological sample solution is 10mg:3mL, described metal heteroatom ZSM-5 zeolite molecular sieve and described H
2O
2The mass volume ratio of solution is 500mg:1mL, and wherein, in Fe-ZSM-5, Fe content is 6%.
Preferably, in step 3, the ultrasonic frequency of described ultrasonic reactor is 40kHz, and ultrasonic power is 12W, and described temperature of reaction is 29～31 ℃, and the described degree of depth is 1.5cm.
Preferably, in step 4, described centrifugal rotational speed is 9500rpm/min, and centrifugation time is 4min.
Preferably, in step 5, described uv-vis spectra scanning wavelength is 200～700nm, and adopting massfraction in described lauryl sodium sulfate-polyacrylamide gel (SDS-PAGE) electrophoretic techniques is 5% concentrated glue, 12.5% separation gel.
Compared with prior art, the present invention has following beneficial effect:
(1) the inventive method adopts metal heteroatom ZSM-5 zeolite molecular sieve, utilizes its avtive spot to be combined with hydrogen peroxide and generates a large amount of hydroxyl radical free radicals, thereby efficient Fenton-like reaction occurs, and can carry out effective pre-treatment to actual biological sample.
(2) the inventive method utilizes Sample Pretreatment Technique with conventional needle, actual biological sample treatment technology to be compared the consumption of having avoided a large amount of severe corrosive acid, base reagent, and handling safety meets " green laboratory " principle.
(3) the inventive method utilizes the heterogeneous Fenton method of ultrasonic supplementary doping metal heteroatom ZSM-5 zeolite molecular sieve to be used for the pretreatment technology of biological sample, efficiently, quick and easy easy operating, have good development prospect in the pre-service of biological sample.
(4) the inventive method can be used as blood wastewater in environment is carried out new method easy, efficient processing, can effectively reduce the blood wastewater of animal blood products factory discharge to the pollution of environment, enterprise is produced accomplish " cleaner production ".
Embodiment
The present invention is described in detail below in conjunction with specific embodiment.Following examples will help those skilled in the art further to understand the present invention, but not limit in any form the present invention.Should be pointed out that to those skilled in the art, without departing from the inventive concept of the premise, can also make some distortion and improvement.These all belong to protection scope of the present invention.
Embodiment 1
The present embodiment relates to a kind of different proportion Fe-ZSM-5 zeolite molecular sieve Fenton method sample-pretreating method, and described method comprises the steps:
Step 1 .Fe-ZSM-5(1%Fe content) synthetic method of zeolite molecular sieve: with Na
2SiO
39H
2O is the silicon source, and tetrapropylene ammonium bromide (TPABr) is template, Fe
2(SO
4)
39H
2O is source of iron.At first with Fe
2(SO
4)
39H
2The O stirring and dissolving is at H
2SO
4In solution, then slowly be added dropwise to the good Na of dissolving in advance under vigorous stirring
2SiO
39H
2O solution adds the tetrapropylene ammonium bromide (TPABr) that measures ratio after fully stirring, and stirs; Mole proportioning of last gained presoma is: 60Na
2O:5Fe
2O
3: 60SiO
2: 2080H
2O:2TPABr:50H
2SO
4The gained aqueous precursor gel is sealed under 180 ℃ reacts 96h, be cooled to room temperature, filter, wash to filtrate pH be 7, after drying, with 1 ℃ of min
-1Speed temperature programme to 550 ℃, keep 6h roasting in muffle furnace, namely get Fe-ZSM-5(1%Fe content) zeolite molecular sieve.
Step 2. accurately take 10mg(with electronic analytical balance and be accurate to 0.0001g) Fe-ZSM-5(1%Fe content) zeolite molecular sieve is in three 5mL polypropylene centrifuge tubes, be numbered B1, H1, F1, add wherein respectively bovine serum albumin(BSA) (BSA), haemoglobin (Hb) and CF (FIB) solution of 3mL0.03mmol/L pH=3 with pipettor, and add according to this 20 μ L30%H
2O
2Solution.
Step 3. sample hose is transferred in 29～31 ℃ of waters bath with thermostatic control, add wherein ice bag and change so that temperature constant in course of reaction in good time, the ultrasonic reactor probe is inserted into 1.5cm under liquid level, ultrasound condition is for (ultrasonic frequency is 30kHz, ultrasonic power is 12W) under, after carrying out ultrasonic reaction 3min, stop reaction.After standing 1min, reopen ultrasonic reactor, after giving ultrasonic reaction 3min, again stop reaction, the discontinuous reaction is carried out stopping after 15min so repeatedly.
Step 4. after pipetting respectively reaction with pipettor, sample solution 400 μ L in the 1.5mL polypropylene centrifuge tube, after the centrifugal 4min of 9500r, pipette supernatant 300 μ L in the 5mL polypropylene centrifuge tube with pipettor, and are to be measured.
Step 5. utilize ultraviolet-visual spectrometer the sample supernatant soln to be carried out spectral scan in 200～700nm wavelength coverage, find that it has maximum absorption band near the 280nm wavelength, can get according to the maximum absorbance value before and after each proteins react 15min, reaction system reaches more than 85% BSA sample degradation rate, Hb sample degradation rate is reached more than 87%, FIB sample degradation rate is reached more than 85%.Utilize the SDS-PAGE gel electrophoresis technology, can get protein component analysis in step 4 sample supernatant, after reaction, the more blank protein example band of sample solution color obviously shoals, and utilizing Image-Lab software to carry out each protein example degradation rate that quantitative test obtains can be corresponding with the uv scan result.
Embodiment 2
The present embodiment relates to a kind of different proportion Fe-ZSM-5 zeolite molecular sieve Fenton method sample-pretreating method, and described method comprises the steps:
Step 1 .Fe-ZSM-5(6%Fe content) synthetic method of zeolite molecular sieve: with Na
2SiO
39H
2O is the silicon source, and tetrapropylene ammonium bromide (TPABr) is template, Fe
2(SO
4)
39H
2O is source of iron.At first with Fe
2(SO
4)
39H
2The O stirring and dissolving is at H
2SO
4In solution, then slowly be added dropwise to the good Na of dissolving in advance under vigorous stirring
2SiO
39H
2O solution adds the tetrapropylene ammonium bromide (TPABr) that measures ratio after fully stirring, and stirs.Mole proportioning of last gained presoma is: 30Na
2O:5Fe
2O
3: 30SiO
2: 1040H
2O:TPABr:25H
2SO
4The gained aqueous precursor gel is sealed under 180 ℃ reacts 96h, be cooled to room temperature, filter, wash to filtrate pH be 7, after drying, with 1 ℃ of min
-1Speed temperature programme to 550 ℃, keep 6h roasting in muffle furnace, get final product to get Fe-ZSM-5(6%Fe content) zeolite molecular sieve.
Step 2. accurately take 10mg(with electronic analytical balance and be accurate to 0.0001g) Fe-ZSM-5(6%Fe content) and zeolite molecular sieve in three 5mL polypropylene centrifuge tubes, be numbered B1, H1, F1.Add wherein respectively bovine serum albumin(BSA) (BSA), haemoglobin (Hb) and CF (FIB) solution of 3mL0.03mmol/L pH=3 with pipettor, and add according to this 20 μ L30%H
2O
2Solution.
Step 3. sample hose is transferred in 29～31 ℃ of waters bath with thermostatic control, add wherein ice bag and change so that temperature constant in course of reaction in good time, the ultrasonic reactor probe is inserted into 1.5cm under liquid level, ultrasound condition is for (ultrasonic frequency is 30kHz, ultrasonic power is 12W) under, after carrying out ultrasonic reaction 3min, stop reaction.After standing 1min, reopen ultrasonic reactor, after giving ultrasonic reaction 3min, again stop reaction, the discontinuous reaction is carried out stopping after 15min so repeatedly.
Step 4. after pipetting respectively reaction with pipettor, sample solution 400 μ L in the 1.5mL polypropylene centrifuge tube, after the centrifugal 4min of 9500r, pipette supernatant 300 μ L in the 5mL polypropylene centrifuge tube with pipettor, and are to be measured.
Step 5. utilize uv-vis spectra the sample supernatant soln to be carried out spectral scan in 200～700nm wavelength coverage, find that it has maximum absorption band near the 280nm wavelength, can get according to the maximum absorbance value before and after each proteins react 15min, reaction system all reaches more than 90% each protein example degradation rate.Utilize the SDS-PAGE gel electrophoresis technology, can get protein component analysis in step 4 sample supernatant, after reaction, the more blank protein example band of sample solution color obviously shoals, and utilizing Image-Lab software to carry out each protein example degradation rate that quantitative test obtains can be corresponding with the uv scan result.
Embodiment 3
The present embodiment relates to a kind of different proportion Fe-ZSM-5 zeolite molecular sieve Fenton method sample-pretreating method, and described method comprises the steps:
Step 1 .Fe-ZSM-5(10%Fe content) synthetic method of zeolite molecular sieve: with Na
2SiO
39H
2O is the silicon source, and tetrapropylene ammonium bromide (TPABr) is template, Fe
2(SO
4)
39H
2O is source of iron.At first with Fe
2(SO
4)
39H
2The O stirring and dissolving is at H
2SO
4In solution, then slowly be added dropwise to the good Na of dissolving in advance under vigorous stirring
2SiO
39H
2O solution adds the tetrapropylene ammonium bromide (TPABr) that measures ratio after fully stirring, and stirs.Mole proportioning of last gained presoma is: 60Na
2O:25Fe
2O
3: 60SiO
2: 2080H
2O:2TPABr:50H
2SO
4The gained aqueous precursor gel is sealed under 180 ℃ reacts 96h, be cooled to room temperature, filter, wash to filtrate pH be 7, after drying, with 1 ℃ of min
-1Speed temperature programme to 550 ℃, keep 6h roasting in muffle furnace, namely get Fe-ZSM-5(10%Fe content) zeolite molecular sieve;
Step 2. accurately take 10mg(with electronic analytical balance and be accurate to 0.0001g) Fe-ZSM-5(10%Fe content) and zeolite molecular sieve in three 5mL polypropylene centrifuge tubes, be numbered B1, H1, F1.Add wherein respectively bovine serum albumin(BSA) (BSA), haemoglobin (Hb) and CF (FIB) solution of 3mL0.03mmol/L pH=3 with pipettor, and add according to this 20 μ L30%H
2O
2Solution.
Step 3. sample hose is transferred in 29～31 ℃ of waters bath with thermostatic control, add wherein ice bag and change so that temperature constant in course of reaction in good time, the ultrasonic reactor probe is inserted into 1.5cm under liquid level, ultrasound condition is for (ultrasonic frequency is 30kHz, ultrasonic power is 12W) under, after carrying out ultrasonic reaction 3min, stop reaction.After standing 1min, reopen ultrasonic reactor, after giving ultrasonic reaction 3min, again stop reaction, the discontinuous reaction is carried out stopping after 15min so repeatedly.
Step 4. after pipetting respectively reaction with pipettor, sample solution 400 μ L in the 1.5mL polypropylene centrifuge tube, after the centrifugal 4min of 9500r, pipette supernatant 300 μ L in the 5mL polypropylene centrifuge tube with pipettor, and are to be measured.
Step 5. utilize uv-vis spectra the sample supernatant soln to be carried out spectral scan in 200～700nm wavelength coverage, find that it has maximum absorption band near the 280nm wavelength, can get according to the maximum absorbance value before and after each proteins react 15min, reaction system reaches more than 90% BSA sample degradation rate, Hb sample degradation rate is reached more than 87%, FIB sample degradation rate is reached more than 90%.Utilize the SDS-PAGE gel electrophoresis technology, can get protein component analysis in step 4 sample supernatant, after reaction, the more blank protein example band of sample solution color obviously shoals, and utilizing Image-Lab software to carry out each protein example degradation rate that quantitative test obtains can be corresponding with the uv scan result.
Embodiment 4
The present embodiment relates to a kind of Fe-ZSM-5 zeolite molecular sieve Fenton method sample-pretreating method, and described method comprises the steps:
Step 1 .Fe-ZSM-5(6%Fe content) synthetic method of zeolite molecular sieve is referring to embodiment 2.
Step 2. accurately take 20mg(with electronic analytical balance and be accurate to 0.0001g) Fe-ZSM-5(6%Fe content in step 1) zeolite molecular sieve is in the 5mL polypropylene centrifuge tube.Add wherein 3mL to dilute by volume 20 times with pipettor and transfer to pH=3 blood sample and 40 μ L30%H2O2 solution.
Step 3. sample hose is transferred in 29～31 ℃ of waters bath with thermostatic control, add wherein ice bag and change so that temperature constant in course of reaction in good time, the ultrasonic reactor probe is inserted into 1.5cm under liquid level, ultrasound condition is for (ultrasonic frequency is 30kHz, ultrasonic power is 12W) under, after carrying out ultrasonic reaction 3min, stop reaction.After standing 1min, reopen ultrasonic reactor, after giving ultrasonic reaction 3min, again stop reaction, the discontinuous reaction is carried out stopping after 15min so repeatedly.
Step 4. after pipetting respectively reaction with pipettor, sample solution 400 μ L in the 1.5mL polypropylene centrifuge tube, after the centrifugal 4min of 9500r, pipette supernatant 300 μ L in the 5mL polypropylene centrifuge tube with pipettor, and are to be measured.
Step 5. utilize the SDS-PAGE gel electrophoresis technology, protein component in step 4 sample supernatant is analyzed, as seen after the reaction, sample solution shoals than blank sample band color, and quantitative test can get according to Image-Lab software, and after reaction, the sample degradation rate reaches more than 90%.Utilize By Hydride Generation-atomic Fluorescence Spectrometry that arsenic content in blood is measured, and add 100 μ g/L arsenic standard solutions to carry out recovery of standard addition mensuration, reaction system is 99.1% to the recovery of As, meets the mensuration requirement.
Embodiment 5
The present embodiment relates to Fe-ZSM-5 zeolite molecular sieve Fenton method sample-pretreating method under a kind of different condition, and described method comprises the steps:
Step 1 .Fe-ZSM-5(6%Fe content) synthetic method of zeolite molecular sieve is referring to embodiment 2.
Step 2. accurately take 2mg(with electronic analytical balance and be accurate to 0.0001g) Fe-ZSM-5(6%Fe content) zeolite molecular sieve is in the 5mL polypropylene centrifuge tube.Bovine serum albumin(BSA) (BSA) solution and the 20 μ L30%H that add wherein 3mL0.03mmol/L pH=3 with pipettor
2O
2Solution.
Step 3. sample hose is transferred in 29～31 ℃ of waters bath with thermostatic control, add wherein ice bag and change so that temperature constant in course of reaction in good time, the ultrasonic reactor probe is inserted into 1.5cm under liquid level, ultrasound condition is for (ultrasonic frequency is 30kHz, ultrasonic power is 12W) under, after carrying out ultrasonic reaction 3min, stop reaction.After standing 1min, reopen ultrasonic reactor, after giving ultrasonic reaction 3min, again stop reaction, the discontinuous reaction is carried out stopping after 15min so repeatedly.
Step 4. after pipetting respectively reaction with pipettor, sample solution 400 μ L in the 1.5mL polypropylene centrifuge tube, after the centrifugal 4min of 9500r, pipette supernatant 300 μ L in the 5mL polypropylene centrifuge tube with pipettor, and are to be measured.
Step 5. utilize uv-vis spectra the sample supernatant soln to be carried out spectral scan in 200～700nm wavelength coverage, find that it has maximum absorption band near the 280nm wavelength, can get according to the maximum absorbance value before and after bovine serum albumin(BSA) reaction 15min, reaction system only has 65% to the protein example degradation rate.Utilize the SDS-PAGE gel electrophoresis technology, can get protein component analysis in step 4 sample supernatant, after reaction, the more blank protein example band of sample solution color slightly shoals, and utilizing Image-Lab software to carry out sample degradation rate that quantitative test obtains can be corresponding with the uv scan result.
Embodiment 6
The present embodiment relates to Fe-ZSM-5 zeolite molecular sieve Fenton method sample-pretreating method under a kind of different condition, and described method comprises the steps:
Step 1 .Fe-ZSM-5(6%Fe content) synthetic method of zeolite molecular sieve is referring to embodiment 2.
Step 2. accurately take 10mg(with electronic analytical balance and be accurate to 0.0001g) Fe-ZSM-5(6%Fe content) zeolite molecular sieve is in the 5mL polypropylene centrifuge tube.Bovine serum albumin(BSA) (BSA) solution and the 20 μ L30%H2O2 solution that add wherein 3mL0.03mmol/L pH=3 with pipettor.
Step 3. sample hose is transferred in 29～31 ℃ of waters bath with thermostatic control, add wherein ice bag and change so that temperature constant in course of reaction in good time, the ultrasonic reactor probe is inserted into 1.5cm under liquid level, ultrasound condition is for (ultrasonic frequency is 30kHz, ultrasonic power is 12W) under, after carrying out ultrasonic reaction 3min, stop reaction.After standing 1min, reopen ultrasonic reactor, after giving ultrasonic reaction 3min, again stop reaction, the discontinuous reaction is carried out stopping after 15min so repeatedly.
Step 4. after pipetting respectively reaction with pipettor, sample solution 400 μ L in the 1.5mL polypropylene centrifuge tube, after the centrifugal 4min of 9500r, pipette supernatant 300 μ L in the 5mL polypropylene centrifuge tube with pipettor, and are to be measured.
Step 5. utilize uv-vis spectra the sample supernatant soln to be carried out spectral scan in 200～700nm wavelength coverage, find that it has maximum absorption band near the 280nm wavelength, can get according to the maximum absorbance value before and after bovine serum albumin(BSA) reaction 15min, reaction system reaches 90% to the protein example degradation rate.Utilize the SDS-PAGE gel electrophoresis technology, can get protein component analysis in step 4 sample supernatant, after reaction, the more blank protein example band of sample solution color slightly shoals, and utilizing Image-Lab software to carry out sample degradation rate that quantitative test obtains can be corresponding with the uv scan result.
Embodiment 7
The present embodiment relates to Fe-ZSM-5 zeolite molecular sieve Fenton method sample-pretreating method under a kind of different condition, and described method comprises the steps:
Step 1 .Fe-ZSM-5(6%Fe content) synthetic method of zeolite molecular sieve is referring to embodiment 2.
Step 2. accurately take 20mg(with electronic analytical balance and be accurate to 0.0001g) Fe-ZSM-5(6%Fe content) zeolite molecular sieve is in the 5mL polypropylene centrifuge tube.Bovine serum albumin(BSA) (BSA) solution and the 20 μ L30%H that add wherein 3mL0.03mmol/L pH=3 with pipettor
2O
2Solution.
Step 3. sample hose is transferred in 29～31 ℃ of waters bath with thermostatic control, add wherein ice bag and change so that temperature constant in course of reaction in good time, the ultrasonic reactor probe is inserted into 1.5cm under liquid level, ultrasound condition is for (ultrasonic frequency is 30kHz, ultrasonic power is 12W) under, after carrying out ultrasonic reaction 3min, stop reaction.After standing 1min, reopen ultrasonic reactor, after giving ultrasonic reaction 3min, again stop reaction, the discontinuous reaction is carried out stopping after 15min so repeatedly.
Step 4. after pipetting respectively reaction with pipettor, sample solution 400 μ L in the 1.5mL polypropylene centrifuge tube, after the centrifugal 4min of 9500r, pipette supernatant 300 μ L in the 5mL polypropylene centrifuge tube with pipettor, and are to be measured.
Step 5. utilize uv-vis spectra the sample supernatant soln to be carried out spectral scan in 200～700nm wavelength coverage, find that it has maximum absorption band near the 280nm wavelength, can get according to the maximum absorbance value before and after bovine serum albumin(BSA) reaction 15min, reaction system reaches 87% to the protein example degradation rate.Utilize the SDS-PAGE gel electrophoresis technology, can get protein component analysis in step 4 sample supernatant, after reaction, the more blank protein example band of sample solution color slightly shoals, and utilizing Image-Lab software to carry out sample degradation rate that quantitative test obtains can be corresponding with the uv scan result.
Embodiment 8
The present embodiment relates to Fe-ZSM-5 zeolite molecular sieve Fenton method sample-pretreating method under a kind of different condition, and described method comprises the steps:
Step 1 .Fe-ZSM-5(6%Fe content) synthetic method of zeolite molecular sieve is referring to embodiment 2.
Step 2. accurately take 10mg(with electronic analytical balance and be accurate to 0.0001g) Fe-ZSM-5(6%Fe content) zeolite molecular sieve is in the 5mL polypropylene centrifuge tube.Bovine serum albumin(BSA) (BSA) solution and the 20 μ L30%H that add wherein 1mL0.03mmol/L pH=3 with pipettor
2O
2Solution.
Step 3. sample hose is transferred in 29～31 ℃ of waters bath with thermostatic control, add wherein ice bag and change so that temperature constant in course of reaction in good time, the ultrasonic reactor probe is inserted into 1.5cm under liquid level, ultrasound condition is for (ultrasonic frequency is 30kHz, ultrasonic power is 12W) under, after carrying out ultrasonic reaction 3min, stop reaction.After standing 1min, reopen ultrasonic reactor, after giving ultrasonic reaction 3min, again stop reaction, the discontinuous reaction is carried out stopping after 15min so repeatedly.
Step 4. after pipetting respectively reaction with pipettor, sample solution 400 μ L in the 1.5mL polypropylene centrifuge tube, after the centrifugal 4min of 9500r, pipette supernatant 300 μ L in the 5mL polypropylene centrifuge tube with pipettor, and are to be measured.
Step 5. utilize uv-vis spectra the sample supernatant soln to be carried out spectral scan in 200～700nm wavelength coverage, find that it has maximum absorption band near the 280nm wavelength, can get according to the maximum absorbance value before and after bovine serum albumin(BSA) reaction 15min, reaction system reaches more than 90% the protein example degradation rate.Utilize the SDS-PAGE gel electrophoresis technology, can get protein component analysis in step 4 sample supernatant, after reaction, the more blank protein example band of sample solution color slightly shoals, and utilizing Image-Lab software to carry out sample degradation rate that quantitative test obtains can be corresponding with the uv scan result.
Embodiment 9
The present embodiment relates to Fe-ZSM-5 zeolite molecular sieve Fenton method sample-pretreating method under a kind of different condition, and described method comprises the steps:
Step 1 .Fe-ZSM-5(6%Fe content) synthetic method of zeolite molecular sieve is referring to embodiment 2.
Step 2. accurately take 10mg(with electronic analytical balance and be accurate to 0.0001g) Fe-ZSM-5(6%Fe content) zeolite molecular sieve is in the 5mL polypropylene centrifuge tube.Bovine serum albumin(BSA) (BSA) solution and the 20 μ L30%H that add wherein 10mL0.03mmol/L pH=3 with pipettor
2O
2Solution.
Step 3. sample hose is transferred in 29～31 ℃ of waters bath with thermostatic control, add wherein ice bag and change so that temperature constant in course of reaction in good time, the ultrasonic reactor probe is inserted into 1.5cm under liquid level, ultrasound condition is for (ultrasonic frequency is 30kHz, ultrasonic power is 12W) under, after carrying out ultrasonic reaction 3min, stop reaction.After standing 1min, reopen ultrasonic reactor, after giving ultrasonic reaction 3min, again stop reaction, the discontinuous reaction is carried out stopping after 15min so repeatedly.
Step 4. after pipetting respectively reaction with pipettor, sample solution 400 μ L in the 1.5mL polypropylene centrifuge tube, after the centrifugal 4min of 9500r, pipette supernatant 300 μ L in the 5mL polypropylene centrifuge tube with pipettor, and are to be measured.
Step 5. utilize uv-vis spectra the sample supernatant soln to be carried out spectral scan in 200～700nm wavelength coverage, find that it has maximum absorption band near the 280nm wavelength, can get according to the maximum absorbance value before and after bovine serum albumin(BSA) reaction 15min, reaction system reaches more than 80% the protein example degradation rate.Utilize the SDS-PAGE gel electrophoresis technology, can get protein component analysis in step 4 sample supernatant, after reaction, the more blank protein example band of sample solution color slightly shoals, and utilizing Image-Lab software to carry out sample degradation rate that quantitative test obtains can be corresponding with the uv scan result.
Embodiment 10
The present embodiment relates to Fe-ZSM-5 zeolite molecular sieve Fenton method sample-pretreating method under a kind of different condition, and described method comprises the steps:
Step 1 .Fe-ZSM-5(6%Fe content) synthetic method of zeolite molecular sieve is referring to embodiment 2.
Step 2. accurately take 10mg(with electronic analytical balance and be accurate to 0.0001g) Fe-ZSM-5(6%Fe content) zeolite molecular sieve is in the 5mL polypropylene centrifuge tube.Bovine serum albumin(BSA) (BSA) solution and the 20 μ L30%H that add wherein 3mL0.03mmol/L pH=2.5 with pipettor
2O
2Solution.
Step 3. sample hose is transferred in 29～31 ℃ of waters bath with thermostatic control, add wherein ice bag and change so that temperature constant in course of reaction in good time, the ultrasonic reactor probe is inserted into 1.5cm under liquid level, ultrasound condition is for (ultrasonic frequency is 30kHz, ultrasonic power is 12W) under, after carrying out ultrasonic reaction 3min, stop reaction.After standing 1min, reopen ultrasonic reactor, after giving ultrasonic reaction 3min, again stop reaction, the discontinuous reaction is carried out stopping after 15min so repeatedly.
Step 4. after pipetting respectively reaction with pipettor, sample solution 400 μ L in the 1.5mL polypropylene centrifuge tube, after the centrifugal 4min of 9500r, pipette supernatant 300 μ L in the 5mL polypropylene centrifuge tube with pipettor, and are to be measured.
Step 5. utilize uv-vis spectra the sample supernatant soln to be carried out spectral scan in 200～700nm wavelength coverage, find that it has maximum absorption band near the 280nm wavelength, can get according to the maximum absorbance value before and after bovine serum albumin(BSA) reaction 15min, reaction system reaches more than 85% the protein example degradation rate.Utilize the SDS-PAGE gel electrophoresis technology, can get protein component analysis in step 4 sample supernatant, after reaction, the more blank protein example band of sample solution color slightly shoals, and utilizing Image-Lab software to carry out sample degradation rate that quantitative test obtains can be corresponding with the uv scan result.
Embodiment 11
The present embodiment relates to Fe-ZSM-5 zeolite molecular sieve Fenton method sample-pretreating method under a kind of different condition, and described method comprises the steps:
Step 1 .Fe-ZSM-5(6%Fe content) synthetic method of zeolite molecular sieve is referring to embodiment 2.
Step 2. accurately take 10mg(with electronic analytical balance and be accurate to 0.0001g) Fe-ZSM-5(6%Fe content) zeolite molecular sieve is in the 5mL polypropylene centrifuge tube.Bovine serum albumin(BSA) (BSA) solution and the 20 μ L30%H that add wherein 3mL0.03mmol/L pH=7.5 with pipettor
2O
2Solution.
Step 3. sample hose is transferred in 29～31 ℃ of waters bath with thermostatic control, add wherein ice bag and change so that temperature constant in course of reaction in good time, the ultrasonic reactor probe is inserted into 1.5cm under liquid level, ultrasound condition is for (ultrasonic frequency is 30kHz, ultrasonic power is 12W) under, after carrying out ultrasonic reaction 3min, stop reaction.After standing 1min, reopen ultrasonic reactor, after giving ultrasonic reaction 3min, again stop reaction, the discontinuous reaction is carried out stopping after 15min so repeatedly.
Step 4. after pipetting respectively reaction with pipettor, sample solution 400 μ L in the 1.5mL polypropylene centrifuge tube, after the centrifugal 4min of 9500r, pipette supernatant 300 μ L in the 5mL polypropylene centrifuge tube with pipettor, and are to be measured.
Step 5. utilize uv-vis spectra the sample supernatant soln to be carried out spectral scan in 200～700nm wavelength coverage, find that it has maximum absorption band near the 280nm wavelength, can get according to the maximum absorbance value before and after bovine serum albumin(BSA) reaction 15min, reaction system reaches more than 85% the protein example degradation rate.Utilize the SDS-PAGE gel electrophoresis technology, can get protein component analysis in step 4 sample supernatant, after reaction, the more blank protein example band of sample solution color slightly shoals, and utilizing Image-Lab software to carry out sample degradation rate that quantitative test obtains can be corresponding with the uv scan result.
Embodiment 12
The present embodiment relates to a kind of Cu-Fe-ZSM-5 zeolite molecular sieve Fenton method sample-pretreating method, and described method comprises the steps:
The synthetic method of step 1 .Cu-Fe-ZSM-5 zeolite molecular sieve: with Na
2SiO
39H
2O is the silicon source, and tetrapropylene ammonium bromide (TPABr) is template, Fe
2(SO
4)
39H
2O is source of iron, CuSO
45H
2O is the copper source.At first with Fe
2(SO
4)
39H
2O, CuSO
45H
2O is by measuring than stirring and dissolving at H
2SO
4In solution, then slowly be added dropwise to the good Na of dissolving in advance under vigorous stirring
2SiO
39H
2O solution adds the tetrapropylene ammonium bromide (TPABr) that measures ratio after fully stirring, and stirs.Mole proportioning of last gained presoma is: 30Na
2O:2Fe
2O
3: 6CuO:30SiO
2: 1040H
2O:TPABr:25H
2SO
4The gained aqueous precursor gel is sealed under 180 ℃ reacts 96h, be cooled to room temperature, filter, wash to filtrate pH be 7, after drying, with 1 ℃ of min
-1Speed temperature programme to 550 ℃, keep 6h roasting in muffle furnace standby, namely get the Cu-Fe-ZSM-5 zeolite molecular sieve;
Step 2. accurately take 10mg(with electronic analytical balance and be accurate to 0.0001g) the Cu-Fe-ZSM-5 zeolite molecular sieve in three 5mL polypropylene centrifuge tubes, be numbered B1, H1, F1.Add wherein respectively bovine serum albumin(BSA) (BSA), haemoglobin (Hb) and CF (FIB) solution of 3mL0.03mmol/L pH=3 with pipettor, and add according to this 20 μ L30%H
2O
2Solution.
Step 3. sample hose is transferred in 29～31 ℃ of waters bath with thermostatic control, add wherein ice bag and change so that temperature constant in course of reaction in good time, the ultrasonic reactor probe is inserted into 1.5cm under liquid level, ultrasound condition is for (ultrasonic frequency is 30kHz, ultrasonic power is 12W) under, after carrying out ultrasonic reaction 3min, stop reaction.After standing 1min, reopen ultrasonic reactor, after giving ultrasonic reaction 3min, again stop reaction, the discontinuous reaction is carried out stopping after 15min so repeatedly.
Step 4. after pipetting respectively reaction with pipettor, sample solution 400 μ L in the 1.5mL polypropylene centrifuge tube, after the centrifugal 4min of 9500r, pipette supernatant 300 μ L in the 5mL polypropylene centrifuge tube with pipettor, and are to be measured.
Step 5. utilize uv-vis spectra the sample supernatant soln to be carried out spectral scan in 200～700nm wavelength coverage, find that it has maximum absorption band near the 280nm wavelength, can get according to the maximum absorbance value before and after each proteins react 15min, reaction system reaches more than 92% BSA sample degradation rate, Hb sample degradation rate is reached more than 95%, FIB sample degradation rate is reached more than 95%.Utilize the SDS-PAGE gel electrophoresis technology, can get protein component analysis in step 4 sample supernatant, after reaction, the more blank protein example band of sample solution color obviously shoals, and utilizing Image-Lab software to carry out each protein example degradation rate that quantitative test obtains can be corresponding with the uv scan result.
Embodiment 13
The present embodiment relates to a kind of Cu-Fe-ZSM-5 zeolite molecular sieve Fenton method sample-pretreating method, and described method comprises the steps:
The synthetic method of step 1 .Cu-Fe-ZSM-5 zeolite molecular sieve is referring to embodiment 12.
Step 2. accurately take 20mg(with electronic analytical balance and be accurate to 0.0001g) the Cu-Fe-ZSM-5 zeolite molecular sieve is in the 5mL polypropylene centrifuge tube.Add wherein 3mL to dilute by volume 20 times with pipettor and transfer to pH=3 blood sample and 40 μ L30%H
2O
2Solution.
Step 3. sample hose is transferred in 29～31 ℃ of waters bath with thermostatic control, add wherein ice bag and change so that temperature constant in course of reaction in good time, the ultrasonic reactor probe is inserted into 1.5cm under liquid level, ultrasound condition is for (ultrasonic frequency is 30kHz, ultrasonic power is 12W) under, after carrying out ultrasonic reaction 3min, stop reaction.After standing 1min, reopen ultrasonic reactor, after giving ultrasonic reaction 3min, again stop reaction, the discontinuous reaction is carried out stopping after 15min so repeatedly.
Step 4. after pipetting respectively reaction with pipettor, sample solution 400 μ L in the 1.5mL polypropylene centrifuge tube, after the centrifugal 4min of 9500r, pipette supernatant 300 μ L in the 5mL polypropylene centrifuge tube with pipettor, and are to be measured.
Step 5. utilize the SDS-PAGE gel electrophoresis technology, protein component in step 4 sample supernatant is analyzed, as seen after the reaction, sample solution shoals than blank sample solution band color, and quantitative test can get according to Image-Lab software, and after reaction, the sample degradation rate reaches more than 90%.Utilize By Hydride Generation-atomic Fluorescence Spectrometry that arsenic content in blood is measured, and add 100 μ g/L arsenic standard solutions to carry out recovery of standard addition mensuration, reaction system reaches 96.7% to the recovery of As, meets the mensuration requirement.
Embodiment 14
The present embodiment relates to a kind of Mn-Fe-ZSM-5 zeolite molecular sieve Fenton method sample-pretreating method, and described method comprises the steps:
The synthetic method of step 1 .Mn-Fe-ZSM-5 zeolite molecular sieve: with Na
2SiO
39H
2O is the silicon source, and tetrapropylene ammonium bromide (TPABr) is template, Fe
2(SO
4)
39H
2O is source of iron, MnSO
44H
2O is the manganese source.At first with Fe
2(SO
4)
39H
2O, MnSO
44H
2O is by measuring than stirring and dissolving at H
2SO
4In solution, then slowly be added dropwise to the good Na of dissolving in advance under vigorous stirring
2SiO
39H
2O solution adds the tetrapropylene ammonium bromide (TPABr) that measures ratio after fully stirring, and stirs.Mole proportioning of last gained presoma is: 30Na
2O:2Fe
2O
3: 6MnO:30SiO
2: 1040H
2O:TPABr:25H
2SO
4The gained aqueous precursor gel is sealed under 180 ℃ reacts 96h, be cooled to room temperature, filter, wash to filtrate pH be 7, after drying, with 1 ℃ of min
-1Speed temperature programme to 550 ℃, keep 6h roasting in muffle furnace, namely get the Mn-Fe-ZSM-5 zeolite molecular sieve;
Step 2. accurately take 10mg(with electronic analytical balance and be accurate to 0.0001g) the Mn-Fe-ZSM-5 zeolite molecular sieve in three 5mL polypropylene centrifuge tubes, be numbered B1, H1, F1.Add wherein respectively bovine serum albumin(BSA) (BSA), haemoglobin (Hb) and CF (FIB) solution of 3mL0.03mmol/L pH=3 with pipettor, and add according to this 20 μ L30%H
2O
2Solution.
Step 3. sample hose is transferred in 29～31 ℃ of waters bath with thermostatic control, add wherein ice bag and change so that temperature constant in course of reaction in good time, the ultrasonic reactor probe is inserted into 1.5cm under liquid level, ultrasound condition is for (ultrasonic frequency is 30kHz, ultrasonic power is 12W) under, after carrying out ultrasonic reaction 3min, stop reaction.After standing 1min, reopen ultrasonic reactor, after giving ultrasonic reaction 3min, again stop reaction, the discontinuous reaction is carried out stopping after 15min so repeatedly.
Step 4. after pipetting respectively reaction with pipettor, sample solution 400 μ L in the 1.5mL polypropylene centrifuge tube, after the centrifugal 4min of 9500r, pipette supernatant 300 μ L in the 5mL polypropylene centrifuge tube with pipettor, and are to be measured.
Step 5. utilize uv-vis spectra the sample supernatant soln to be carried out spectral scan in 200～700nm wavelength coverage, find that it has maximum absorption band near the 280nm wavelength, can get according to the maximum absorbance value before and after each proteins react 15min, reaction system reaches more than 87% BSA sample degradation rate, Hb sample degradation rate is reached more than 90%, FIB sample degradation rate is reached more than 90%.Utilize the SDS-PAGE gel electrophoresis technology, can get protein component analysis in step 4 sample supernatant, after reaction, the more blank protein example band of sample solution color obviously shoals, and utilizing Image-Lab software to carry out each protein example degradation rate that quantitative test obtains can be corresponding with the uv scan result.
Embodiment 15
The present embodiment relates to a kind of Mn-Fe-ZSM-5 zeolite molecular sieve Fenton method sample-pretreating method, and described method comprises the steps:
The synthetic method of step 1 .Mn-Fe-ZSM-5 zeolite molecular sieve is referring to embodiment 14.
Step 2. accurately take 18.6mg(with electronic analytical balance and be accurate to 0.0001g) the Mn-Fe-ZSM-5 zeolite molecular sieve is in the 5mL polypropylene centrifuge tube.Add wherein 3mL to dilute by volume 20 times with pipettor and transfer to pH=3 blood sample and 40 μ L30%H
2O
2Solution.
Step 3. sample hose is transferred in 29～31 ℃ of waters bath with thermostatic control, add wherein ice bag and change so that temperature constant in course of reaction in good time, the ultrasonic reactor probe is inserted into 1.5cm under liquid level, ultrasound condition is for (ultrasonic frequency is 30kHz, ultrasonic power is 12W) under, after carrying out ultrasonic reaction 3min, stop reaction.After standing 1min, reopen ultrasonic reactor, after giving ultrasonic reaction 3min, again stop reaction, the discontinuous reaction is carried out stopping after 15min so repeatedly.
Step 4. after pipetting respectively reaction with pipettor, sample solution 400 μ L in the 1.5mL polypropylene centrifuge tube, after the centrifugal 4min of 9500r, pipette supernatant 300 μ L in the 5mL polypropylene centrifuge tube with pipettor, and are to be measured.
Step 5. utilize the SDS-PAGE gel electrophoresis technology, protein component in step 4 sample supernatant is analyzed, as seen after the reaction, sample solution shoals than blank sample solution band color, and quantitative test can get according to Image-Lab software, and after reaction, the sample degradation rate reaches more than 90%.Utilize By Hydride Generation-atomic Fluorescence Spectrometry that arsenic content in blood is measured, and add 100 μ g/L arsenic standard solutions to carry out recovery of standard addition mensuration, reaction system is 100.1% to the recovery of As, meets the mensuration requirement.
Embodiment 16
The present embodiment relates to a kind of Ti-Fe-ZSM-5 zeolite molecular sieve Fenton method sample-pretreating method, and described method comprises the steps:
The synthetic method of step 1 .Ti-Fe-ZSM-5 zeolite molecular sieve: with Na
2SiO
39H
2O is the silicon source, and tetrapropylene ammonium bromide (TPABr) is template, Fe
2(SO
4)
39H
2O is source of iron, Ti (SO
4)
2Be the titanium source.At first with Fe
2(SO
4)
39H
2O, Ti (SO
4)
2By measuring than stirring and dissolving at H
2SO
4In solution, then slowly be added dropwise to the good Na of dissolving in advance under vigorous stirring
2SiO
39H
2O solution adds the tetrapropylene ammonium bromide (TPABr) that measures ratio after fully stirring, and stirs.Mole proportioning of last gained presoma is: 30Na
2O:2Fe
2O
3: 6TiO
2: 30SiO
2: 1040H
2O:TPABr:25H
2SO
4The gained aqueous precursor gel is sealed under 180 ℃ reacts 96h, be cooled to room temperature, filter, wash to filtrate pH be 7, after drying, with 1 ℃ of min
-1Speed temperature programme to 550 ℃, keep 6h roasting in muffle furnace, namely get the Ti-Fe-ZSM-5 zeolite molecular sieve;
Step 2. accurately take 10mg(with electronic analytical balance and be accurate to 0.0001g) the Ti-Fe-ZSM-5 zeolite molecular sieve in three 5mL polypropylene centrifuge tubes, be numbered B1, H1, F1.Add wherein respectively bovine serum albumin(BSA) (BSA), haemoglobin (Hb) and CF (FIB) solution of 3mL0.03mmol/L pH=3 with pipettor, and add according to this 20 μ L30%H
2O
2Solution.
Step 3. sample hose is transferred in 29～31 ℃ of waters bath with thermostatic control, add wherein ice bag and change so that temperature constant in course of reaction in good time, the ultrasonic reactor probe is inserted into 1.5cm under liquid level, ultrasound condition is for (ultrasonic frequency is 30kHz, ultrasonic power is 12W) under, after carrying out ultrasonic reaction 3min, stop reaction.After standing 1min, reopen ultrasonic reactor, after giving ultrasonic reaction 3min, again stop reaction, the discontinuous reaction is carried out stopping after 15min so repeatedly.
Step 4. after pipetting respectively reaction with pipettor, sample solution 400 μ L in the 1.5mL polypropylene centrifuge tube, after the centrifugal 4min of 9500r, pipette supernatant 300 μ L in the 5mL polypropylene centrifuge tube with pipettor, and are to be measured.
Step 5. utilize uv-vis spectra the sample supernatant soln to be carried out spectral scan in 200～700nm wavelength coverage, find that it has maximum absorption band near the 280nm wavelength, can get according to the maximum absorbance value before and after each proteins react 15min, reaction system reaches more than 85% BSA sample degradation rate, Hb sample degradation rate is reached more than 90%, FIB sample degradation rate is reached more than 87%.Utilize the SDS-PAGE gel electrophoresis technology, can get protein component analysis in step 4 sample supernatant, after reaction, the more blank protein example band of sample solution color obviously shoals, and utilizing Image-Lab software to carry out each protein example degradation rate that quantitative test obtains can be corresponding with the uv scan result.
Embodiment 17
The present embodiment relates to a kind of Ti-Fe-ZSM-5 zeolite molecular sieve Fenton method sample-pretreating method, and described method comprises the steps:
The synthetic method of step 1 .Ti-Fe-ZSM-5 zeolite molecular sieve is referring to embodiment 16.
Step 2. accurately take 20mg(with electronic analytical balance and be accurate to 0.0001g) the Ti-Fe-ZSM-5 zeolite molecular sieve is in the 5mL polypropylene centrifuge tube.Add wherein 3mL to dilute by volume 20 times with pipettor and transfer to pH=3 blood sample and 40 μ L30%H
2O
2Solution.
Step 3. sample hose is transferred in 29～31 ℃ of waters bath with thermostatic control, add wherein ice bag and change so that temperature constant in course of reaction in good time, the ultrasonic reactor probe is inserted into 1.5cm under liquid level, ultrasound condition is for (ultrasonic frequency is 30kHz, ultrasonic power is 12W) under, after carrying out ultrasonic reaction 3min, stop reaction.After standing 1min, reopen ultrasonic reactor, after giving ultrasonic reaction 3min, again stop reaction, the discontinuous reaction is carried out stopping after 15min so repeatedly.
Step 4. after pipetting respectively reaction with pipettor, sample solution 400 μ L in the 1.5mL polypropylene centrifuge tube, after the centrifugal 4min of 9500r, pipette supernatant 300 μ L in the 5mL polypropylene centrifuge tube with pipettor, and are to be measured.
Step 5. utilize the SDS-PAGE gel electrophoresis technology, protein component in step 4 sample supernatant is analyzed, as seen after the reaction, sample solution shoals than blank sample solution band color, and quantitative test can get according to Image-Lab software, and after reaction, the sample degradation rate reaches more than 90%.Utilize By Hydride Generation-atomic Fluorescence Spectrometry that arsenic content in blood is measured, and add 100 μ g/L arsenic standard solutions to carry out recovery of standard addition mensuration, reaction system is 96.4% to the recovery of As, meets the mensuration requirement.
Implementation result: the metal heteroatom zeolite molecular sieve that the present embodiment 1～17 makes is according to ultraviolet visible spectrometry, SDS-PAGE gel electrophoresis technology, By Hydride Generation-atomic Fluorescence Spectrometry, to adding different proportion Fe atom ZSM-5 type zeolite molecular sieve reaction system that each biological sample degradation efficiency is compared and can get, when doped F e atomic mass mark was 6%, degradation efficiency was the highest; Degradation rate to bovine serum albumin(BSA) under the differential responses condition can get, the top condition of reaction is Fe-ZSM-5(6%Fe content) quality of zeolite molecular sieve and described biological sample solution: volume=10mg:3mL, Fe-ZSM-5(6%Fe content) zeolite molecular sieve and described H
2O
2The quality of solution: volume=500mg:1mL, reacting best potential of hydrogen is pH=3.Degradation efficiency contrast to the different heteroatoms ZSM-5 type zeolite molecular sieve reaction systems of adulterating can get, and adds double heteroatoms Cu-Fe-ZSM-5 type zeolite molecular sieve reaction system the highest to the degradation efficiency of several samples, has significant superiority.Utilize By Hydride Generation-atomic Fluorescence Spectrometry that arsenic content in blood is measured, its recovery all reaches more than 95%, and good testing result is arranged.
In sum: the inventive method adopts metal heteroatom ZSM-5 zeolite molecular sieve, utilize its avtive spot to be combined with hydrogen peroxide and generate a large amount of hydroxyl radical free radicals, thereby efficient Fenton-like reaction occurs, can carry out effective pre-treatment to actual biological sample; Utilize Sample Pretreatment Technique with conventional needle, actual biological sample treatment technology to be combined and compare the consumption of having avoided a large amount of severe corrosive acid, base reagent, handling safety meets " green laboratory " principle; Utilize the heterogeneous Fenton method of ultrasonic supplementary doping metal heteroatom ZSM-5 zeolite molecular sieve for the pretreatment technology of biological sample, efficient, quick and easy easy operating has good development prospect in the pre-service of biological sample; Can be used as blood wastewater in environment is carried out new method easy, efficient processing, can effectively reduce the blood wastewater of animal blood products factory discharge to the pollution of environment, enterprise is produced accomplish " cleaner production ".
The above, it is only preferred embodiment of the present invention, be not that the present invention is done any pro forma restriction, any content that does not break away from technical solution of the present invention, to any simple modification, equivalent variations and modification that above embodiment does, all belong to the scope of technical solution of the present invention according to technical spirit of the present invention.
Claims (12)
1. a metal heteroatom zeolite molecular sieve Fenton method sample-pretreating method, is characterized in that, described method comprises the steps:
Step 1, metal heteroatom ZSM-5 zeolite molecular sieve synthetic; Described metal heteroatom ZSM-5 zeolite molecular sieve is Fe-ZSM-5 zeolite molecular sieve or M-Fe-ZSM-5 zeolite molecular sieve, and wherein M is Cu, Mn or Ti;
Step 2 adds described metal heteroatom ZSM-5 zeolite molecular sieve in centrifuge tube, and adds wherein biological sample solution and H
2O
2Solution;
Step 3 is transferred to centrifuge tube in water bath with thermostatic control, the ultrasonic reactor probe is inserted under liquid level reaction;
Step 4 after question response is completed, is taken out centrifuge tube, and is centrifugal, pipettes supernatant, to be measured;
Step 5 records maximum absorbance before and after proteins react according to ultraviolet-visual spectrometer, utilizes sodium dodecyl sulfate-polyacrylamide gel electrophoresis technology and Image-Lab software to carry out quantitative test and mensuration to the band intensity before and after proteins react.
2. metal heteroatom zeolite molecular sieve Fenton method sample-pretreating method as claimed in claim 1, is characterized in that, the synthetic of described Fe-ZSM-5 zeolite molecular sieve is specially:
At first with Fe
2(SO
4)
39H
2The O stirring and dissolving is at H
2SO
4In solution, then slowly be added dropwise to the good Na of dissolving in advance under vigorous stirring
2SiO
39H
2O solution adds the tetrapropylene ammonium bromide that measures ratio after fully stirring, and stirs, and gets aqueous precursor gel; The gained aqueous precursor gel is sealed under 180 ℃ reacts 96h, through cooling, filtration, after washing, drying, be transferred to roasting under 550 ℃ of conditions in muffle furnace.
3. metal heteroatom zeolite molecular sieve Fenton method sample-pretreating method as claimed in claim 2, is characterized in that, in described Fe-ZSM-5 zeolite molecular sieve, the mass percent of iron is 1～10%.
4. metal heteroatom zeolite molecular sieve Fenton method sample-pretreating method as claimed in claim 1, is characterized in that, the synthetic of described M-Fe-ZSM-5 zeolite molecular sieve is specially:
At first with Fe
2(SO4)
39H
2O, sulfuric acid M salt are by measuring than stirring and dissolving at H
2SO
4In solution, then slowly be added dropwise to the good Na of dissolving in advance under vigorous stirring
2SiO
39H
2O solution adds the tetrapropylene ammonium bromide that measures ratio after fully stirring, and stirs, and gets aqueous precursor gel; The gained aqueous precursor gel is sealed under 180 ℃ reacts 96h, through cooling, filtration, after washing, drying, be transferred to roasting under 550 ℃ of conditions in muffle furnace.
5. as the described metal heteroatom zeolite molecular sieve of claim 1-4 any one Fenton method sample-pretreating method, it is characterized in that, in step 2, described biological sample solution is specially: with contained albumin, haemoglobin, fibrinogenic content ratio in blood, with the formulated protein solution of NaCl solution dissolving, wherein the volumetric molar concentration of NaCl solution is 0.15moL/L respectively.
6. metal heteroatom zeolite molecular sieve Fenton method sample-pretreating method as claimed in claim 5, it is characterized in that, in step 2, the pH value scope of described biological sample solution is 2.5～7.5, and it is 1moL/L HCl and 1moL/L NaOH that the pH of described biological sample solution regulates reagent.
7. as the described metal heteroatom zeolite molecular sieve of claim 1-4 any one Fenton method sample-pretreating method, it is characterized in that, in step 2, described H
2O
2The solution quality percent concentration is 30%.
8. metal heteroatom zeolite molecular sieve Fenton method sample-pretreating method as claimed in claim 1, it is characterized in that, in step 2, the mass volume ratio of described metal heteroatom ZSM-5 zeolite molecular sieve and described biological sample solution is (1～10): 1, and described metal heteroatom ZSM-5 zeolite molecular sieve and described H
2O
2The mass volume ratio of solution is (100～1000): 1.
9. metal heteroatom zeolite molecular sieve Fenton method sample-pretreating method as claimed in claim 8, it is characterized in that, the mass volume ratio of described metal heteroatom ZSM-5 zeolite molecular sieve and described biological sample solution is 10mg:3mL, described metal heteroatom ZSM-5 zeolite molecular sieve and described H
2O
2The mass volume ratio of solution is 500mg:1mL, and wherein, in Fe-ZSM-5, Fe content is 6%.
10. as the described metal heteroatom zeolite molecular sieve of claim 1-4 any one Fenton method sample-pretreating method, it is characterized in that, in step 3, the ultrasonic frequency of described ultrasonic reactor is 40kHz, ultrasonic power is 12W, and described temperature of reaction is 29～31 ℃.
11. as the described metal heteroatom zeolite molecular sieve of claim 1-4 any one Fenton method sample-pretreating method, it is characterized in that, in step 4, described centrifugal rotational speed is 9500rpm/min, centrifugation time is 4min.
12. as the described metal heteroatom zeolite molecular sieve of claim 1-4 any one Fenton method sample-pretreating method, it is characterized in that, in step 5, described uv-vis spectra scanning wavelength is 200～700nm, and adopting massfraction in described sodium dodecyl sulfate-polyacrylamide gel electrophoresis technology is 5.0% concentrated glue, 12.5% separation gel.
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| CN110702844A (en) * | 2019-10-11 | 2020-01-17 | 山西省化工研究所（有限公司） | Method for analyzing and determining aluminum content in organic nucleating agent by Fenton-like reagent |
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Cited By (5)
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| CN108254322A (en) * | 2016-12-28 | 2018-07-06 | 中粮集团有限公司 | Cereal digestion procedure and its application |
| CN108554442A (en) * | 2018-05-15 | 2018-09-21 | 中国石油大学(华东) | Novel ferrimanganic two-component catalyst and its Catalytic processes decomposed for hydrogen peroxide |
| CN108816272A (en) * | 2018-05-15 | 2018-11-16 | 中国石油大学(华东) | A kind of novel ferrimanganic two-component catalyst and catalytic degradation acetone method |
| CN109238949A (en) * | 2018-09-19 | 2019-01-18 | 浙江大学 | A method of micro- plastic density distribution in detection marine organisms soft tissue |
| CN110702844A (en) * | 2019-10-11 | 2020-01-17 | 山西省化工研究所（有限公司） | Method for analyzing and determining aluminum content in organic nucleating agent by Fenton-like reagent |
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| CN103383321B (en) | 2016-05-04 |
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