CN102539606A - Determination method of trivalent aluminum in aquatic product and product thereof - Google Patents

Abstract

The invention discloses a determination method of trivalent aluminum in an aquatic product and a product thereof. The determination method comprises the following steps: extracting trivalent aluminum in the aquatic product and the product thereof by using a mixed liquid of 0.0040mol/L lithium hydroxide, 0.0060mol/L pyridine-2.6-dicarboxylic acid and 0.090mol/L glacial acetic acid; separating in a high performance liquid chromatograph; eluting Al in various forms with a flowing phase sequentially because a cation-exchange chromatographic column has different absorption capabilities on Al in various forms; analyzing and detecting 27AL in an eluent through an inductive coupling plasma spectrometer, thus more complete biological characteristics of Al in the aquatic product are given. According to the invention, the experiment conditions are reasonably selected, and the detection data is accurate and reliable and is not influenced by digestion time and temperature; and by using the method, the contents of trivalent aluminum in the aquatic product and the product thereof can be determined accurately.

Description

The assay method of trivalent aluminium in aquatic products and the goods thereof

Technical field:

The invention belongs to the aquatic products detection technique, relate to a kind of method of measuring trivalent aluminium in aquatic products and the goods thereof with high performance liquid chromatography-inductively coupled plasma mass spectrometer (HPLC-ICP-MS) coupling technique particularly.

Background technology:

Current; A forward position content of elementary analytical chemistry is that the different shape to element carries out qualitative and quantitative; And be not only the total amount of measuring element, this mainly is because the different shape of identity element possibly have diverse chemical property, aluminium is one of them typical element.The chemical form of aluminium element in hydrobionts such as marine alga is different from terrestrial organism, and element morphology difference is to influence the key factor that the marine alga edible safety is estimated, but the research of this respect is less, and particularly relevant marine alga aspect is close to blank.

According to up-to-date discovering, aluminium has variform in physical environment and the living things system, and the toxicity of different shape aluminium differs greatly.Result of study shows that there is variform in aluminium in the aquatic products, mainly with stable, be difficult for being existed by the organo-aluminium form of organism, and higher inorganic states aluminium (trivalent aluminium) content of toxicity is lower.About aquatic products processes such as process washing, the cooking in edible process, whether the form and the content of the aluminium in the body have changed, and the research final conclusion is not arranged as yet yet in addition.

The detection method of existing aluminium is: the assay method of aluminium in the flour-made food of stipulating among the GB GB/T5009.182-2003; And algae such as sea-tangle, laver are to lean on the photosynthesis self-sow in breeding process; Be a kind of ocean food of pure natural; The processing of product such as sea-tangle, laver generally is through washing dish, dehydration, moulding, oven dry, packing, weigh and flow process such as storage; Especially in process, there is no need to add leavening agent aluminium potassium sulfate (potassium alum) and alum (ammonia-alum), the processing of this and Flour product or dilated food is diverse.Therefore, adopt this method to detect marine alga and be the aluminium residual quantity of aquatic products and the goods thereof of main food chain all exceed standard ((mg/kg)≤100 of limiting the quantity of of aluminium among the food additives hygienic standard GB2760-2007) with marine alga.Aluminium in the aquatic products is not to be the aluminium potassium sulfate of GB2760 indication or the trivalent aluminium in the Exsiccated Ammonium Alum fully---and " non-those aluminium of this aluminium ".Therefore to the research of al species analytical approach in the aquatic products with set up extremely urgently, become the important front edge problem of national natural science base growth.As early as possible the assay method of trivalent aluminium is studied, and set up and to be suitable for the standard method that all kinds of aquatic products detect and to be very important.Still there is not the method standard of measuring trivalent aluminium in the aquatic products in present international ISO and the CAC standard.

Summary of the invention:

The technical matters that the present invention will solve provides the assay method of trivalent aluminium in a kind of aquatic products and the goods thereof; Use high performance liquid chromatography-inductively coupled plasma mass spectrometer coupling (HPLC-ICP-MS) morphological analysis technology; Form to its trivalent aluminium is carried out qualitative and quantitative analysis; Accurately measure the trivalent aluminium in aquatic products and the goods thereof, thereby provide biological characteristics more accurately and reliably.

The present invention accomplishes according to following method of operating:

The assay method of trivalent aluminium in a kind of aquatic products and the goods thereof; May further comprise the steps: instrument of 1, selecting for use and equipment, HPLC-ICP-MS combined system, 2, configuration reagent and standard solution; 3, the step of sample preparation, 4, the used instrument condition 5 of experiment, sample detection and result's calculating; Concrete steps are following:

1) instrument of selecting for use and equipment, the HPLC-ICP-MS combined system:

(1), HPLC part: use SERIES200 high-pressure liquid phase pump, be furnished with automatic sampler, be furnished with CG5A 4 * 250mm cation-exchange chromatography post of guard column CG5A 4 * 50mm and identical material filler;

(2), ICP-MS: be furnished with the dynamic response pond, sampling system, radio-frequency signal generator, plasma system, interface, ion lens system, quadrupole rod, vacuum system, detecting device, air-path control system;

(3), cooling circulating water;

(4), constant temperature water bath oscillator;

(5), sand core filter;

(6), circulating vacuum pump;

(7), hydro-extractor;

(8), numerical control ultrasonic cleaning machine;

(9), the water system miillpore filter of 0.45 μ m;

(10), the argon gas of purity >=99.99%;

2) configuration reagent and standard solution:

(1), leaching liquor: the mixed liquor of 0.0040mol/L lithium hydroxide, 0.0060mol/L pyridine-2.6-dicarboxylic acid, 0.090mol/L glacial acetic acid;

(2), moving phase: the mixed liquor of 0.0040mol/L lithium hydroxide, 0.0060mol/L pyridine-2.6-dicarboxylic acid, 0.090mol/L glacial acetic acid, before using through 0.45 μ m membrane filtration and ultrasonic degas;

All reagent all is that analysis is pure or top grade is pure, and ultrapure water is all used in whole experiment; Warp 15% nitric acid dousing 24h before glass apparatus uses;

(3), the standard of AL (III) is used liquid:

AL (III) standard is used liquid: the storing solution that 100 μ g/mL AL (III) standard solution is diluted to 1.00 μ g/mL AL (III); Putting 4 ℃ of refrigerators deposits subsequent use; AL (III) storing solution by 1.00 μ g/mL uses liquid through AL (III) standard that moving phase is diluted to 50.0ng/mL, 100.0ng/mL, 250.0ng/mL, 500.0ng/mL, 1000.0ng/mL, and is existing with join at present;

3) step of sample preparation:

Take by weighing through the dry sample 0.400g～0.800g that pulverized 40 mesh sieves and place 50mL tool plug scale test tube, add extract 20mL mixing, or bright appearance is smashed homogenate to pieces and taken by weighing 2.000g～4.000g and place 50mL tool plug scale test tube; Add extract 20mL mixing, to 70 ℃ of water bath with thermostatic control shaking table 1h, after the abundant lixiviate of sample; Take out cooling, the centrifugal 15min of 4000r/min takes out supernatant with 0.45 micron water system filtering with microporous membrane; Move in the sample introduction bottle, last machine is measured the value of trivalent aluminium;

4) the used instrument condition of experiment:

(1), high-efficient liquid phase chromatogram condition HPLC: moving phase is the mixed liquor of 0.0040mol/L lithium hydroxide, 0.0060mol/L pyridine-2.6-dicarboxylic acid, 0.090mol/L glacial acetic acid, flow velocity 1mL/min; Sampling volume 50～100 μ L, auto injection;

(2), inductively coupled plasma mass spectrometer condition ICP-MS: radio-frequency power 1150～1600W, plasma flow speed 15～20L/min, secondary air speed 1.2～1.5L/min; Sampling time 10min; Measure mass number ²⁷AL;

5) sample detection and result calculate:

(1), qualitative: the start back is provided with by above-mentioned condition, after waiting to stablize, does typical curve earlier, measures the sample measuring liquid of handling well of treating then; Under above-mentioned high performance liquid chromatography and inductively coupled plasma mass spectrometer condition, adopt full-automatic injector sample introduction 50～100 μ L to measure, high performance liquid chromatograph separates, and the inductively coupled plasma mass spectrometer is right ²⁷AL detects; Trivalent aluminium AL (III) retention time is about 3.172min under above-mentioned instrument condition;

(2), quantitative: as to treat that the chromatographic peak retention time of AL in the sample measuring liquid (III) and standard use liquid phase within ± 10%, promptly to think this material to be determined than variation range;

The result calculates and statement

With the external standard method in the data processing software, confirm AL (III) retention time and degree of separation pulse strength, be horizontal ordinate with concentration; The pulse strength integration is an ordinate; The drawing standard working curve then with appearance liquid peak analyzing and processing, can obtain treating the concentration of the AL (III) in the sample measuring liquid; Calculate the content of trivalent aluminium in the derived sample by following formula (1):

X＝C×V/m×1000............... (1)

In the formula:

The content of trivalent aluminium in the X-sample, mg/kg;

C-treats the concentration of trivalent aluminium in the sample measuring liquid, ng/ml;

The m-sample weighting amount, g;

V-measures total liquid volume, mL.

The present invention's beneficial effect compared with prior art is:

1, after the AL (III) in aquatic products and the goods thereof extracts with extract; Separate through high performance liquid chromatograph; Because the cation-exchange chromatography post is different to the adsorptive power of the various forms of Al, moving phase is with its wash-out successively, and eluent is right through the inductively coupled plasma mass spectrometer ²⁷AL carries out analyzing and testing, and this choice of experimental conditions is reasonable, detects data accurately and reliably.

2, the form of AL (III) does not receive extraction time and temperature effect in the inventive method mensuration aquatic products sample and the goods thereof, can accurately measure the content of AL in aquatic products and the goods thereof (III).

3, the present invention has filled up the blank that total aluminium assay method is only arranged in the present GB and AL (III) form detection method is not arranged, the evaluation aquatic products and the Products Quality thereof of ability accurate science.

Description of drawings:

Fig. 1: AL (III) standard is used liquid form spectrogram;

Fig. 2: the sea-tangle sample is lixiviate 0.5～2.0h AL (III) form spectrogram in 70 ℃ of waters bath with thermostatic control;

Embodiment:

Be described in detail technology contents of the present invention through embodiment below:

The present invention is for utilizing the morphological analysis method of high performance liquid chromatography-inductively coupled plasma mass spectrometer coupling (HPLC-ICP-MS) morphological analysis technology to trivalent aluminium in aquatic products and the goods thereof; Step comprises: instrument of 1, selecting for use and equipment; 2, configuration reagent and standard solution; 3, the step of sample extraction, 4, the used instrument condition of experiment, 5, sample detection and result's calculating.

Aquatic products are various in style, like shellfish, fish, shell-fish, siphonopods and various algae etc.; Inrichment and mechanism to aluminium in its growth course have nothing in common with each other; The content and the existence form of aluminium also differ greatly in its body; The chemical form of aluminium element in hydrobionts such as marine alga is different from terrestrial organism; Element morphology difference is to influence the key factor that the marine alga edible safety is estimated, this to be present in the different biosomes with algae in the mensuration of aluminium have higher requirement, especially with algae for very.Therefore, we select the most representative and embodiment of maximum algae as checking that dispute in the aquatic products.

Embodiment 1: the mensuration of AL in the sea-tangle (III)

1, instrument and the equipment selected for use of present embodiment: HPLC-I CP-MS combined system:

(1), HPLC (U.S. Perkin-Elmer company; Model Series 200) part: use SERIES200 high-pressure liquid phase pump; Be furnished with automatic sampler, be furnished with CG5A 4 * 250mm cation-exchange chromatography post of guard column CG5A 4 * 50mm and identical material filler;

(2), ICP-MS (U.S. Perkin-Elmer company, model ELAN DRCII) part: be furnished with dynamic response pond (DRC), sampling system, radio-frequency signal generator; Plasma system, interface, ion lens system, quadrupole rod; Vacuum system, detecting device, air-path control system;

(3), cooling circulating water (U.S. POLYSCIENCE company, model SHA-C);

(4), constant temperature water bath oscillator (Shanghai and glad instrument and meter company limited, model SHA-C);

(5), sand core filter (core filtration unit, model SH/T0093);

(6), circulating vacuum pump (Shanghai Yarong Biochemical Instrument Plant, model SHZ-III);

(7), hydro-extractor (Beijing Lei Boer hydro-extractor company limited, model LD5-10);

(8), numerical control ultrasonic cleaning machine (some company of city of Kunshan's ultrasonic instrument, model KQ-600DE);

(9), water system miillpore filter (0.45 μ m);

(10), argon gas (purity >=99.99%).

2, configuration reagent and standard solution;

(1), leaching liquor takes by weighing lithium hydroxide 1.700g, pyridine-2.6-dicarboxylic acid 1.000g, glacial acetic acid 5.420g; Slowly be settled to 1000mL with ultrapure water, obtain the mixed liquor of 0.0040mol/L lithium hydroxide, 0.0060mol/L pyridine-2.6-dicarboxylic acid, 0.090mol/L glacial acetic acid behind the mixing.

(2), moving phase: the mixed liquor of 0.0040mol/L lithium hydroxide, 0.0060mol/L pyridine-2.6-dicarboxylic acid, 0.090mol/L glacial acetic acid.Use preceding through 0.45 μ m membrane filtration and ultrasonic degas.

All reagent all is that analysis is pure or top grade is pure, and ultrapure water is all used in whole experiment; Warp 15% nitric acid dousing 24h before glass apparatus uses;

(3), the standard of AL (III) is used liquid:

AL (III) standard is used liquid: buy the standard solution of 100 μ g/mLAL (III) from national standard material center, be diluted to the storing solution (put 4 ℃ refrigerators deposit subsequent use) of 1.00 μ g/mL as AL (III); 1.00 the AL of μ g/mL (III) storing solution uses liquid through AL (III) standard that moving phase is diluted to 50.0ng/mL, 100.0ng/mL, 250.0ng/mL, 500.0ng/mL, 1000.0ng/mL, and is existing with join at present.

3, the step of sample preparation:

Take by weighing through the about 0.400g of the dry sample of pulverizing 40 mesh sieves and place 50mL tool plug scale test tube,,, after the abundant lixiviate of sample, take out cooling to 70 ℃ of water bath with thermostatic control shaking table 1h with leaching liquor 20mL mixing.The centrifugal 15min of 4000r/min takes out supernatant with 0.45 micron water system filtering with microporous membrane, moves in the sample introduction bottle value of last machine mensuration trivalent aluminium.

4, the used instrument condition of experiment;

(1), high-efficient liquid phase chromatogram condition HPLC: the mixed liquor of moving phase 0.0040mol/L lithium hydroxide, 0.0060mol/L pyridine-2.6-dicarboxylic acid, 0.090mol/L glacial acetic acid, flow velocity 1mL/min; Sampling volume 50 μ L, auto injection.

(2), inductively coupled plasma mass spectrometer condition ICP-MS: radio-frequency power 1150W, plasma flow speed 15L/min, secondary air speed 1.2L/min; Sampling time 10min.Measure mass number ²⁷AL.

5, sample detection and result calculate:

(1), qualitative: the start back is provided with by above-mentioned condition, after waiting to stablize, does typical curve earlier, measures the sample solution of handling well then.Under above-mentioned high performance liquid chromatography and inductively coupled plasma mass spectrometer condition, adopt full-automatic injector sample introduction 50 μ L to measure, high performance liquid chromatograph separates, and the inductively coupled plasma mass spectrometer is right ²⁷AL detects; Trivalent aluminium AL (III) retention time is about 3.172min under above-mentioned instrument condition; See Fig. 1 for details;

(2), quantitative: the chromatographic peak retention time of AL in the testing sample (III) uses liquid phase within ± 10%, promptly to think this material to be determined than variation range with standard;

The result calculates and statement

With the external standard method in the data processing software (chromera), confirm AL (III) retention time and degree of separation pulse strength, with concentration horizontal ordinate; The pulse strength integration is an ordinate, and the drawing standard working curve is preserved standard working curve; Then with sample peak analyzing and processing; Can obtain treating the concentration of the AL (III) in the sample measuring liquid, calculate, can get the content of trivalent aluminium in the sample by following formula (1):

X＝C×V/m×1000............... (1)

In the formula:

The content of trivalent aluminium in the X-sample, mg/kg;

C-treats the concentration of AL in the sample measuring liquid (III), ng/ml;

The m-sample weighting amount, g;

V-measures total liquid volume, mL;

6, result and discussion:

The analytical performance of table 2, instrument

Can be known by table 2: the analytical performance of this instrument meets fully measures Al (III);

The mensuration result of table 3, sea-tangle recovery of standard addition (in butt):

Can be known by table 3: we add target Al (III) recovery 88.7%, and recovering effect is desirable.

For our measure taked accurately and reliably of guaranteeing experimental data has: 1. sample need be pulverized 40 mesh sieves and can make sample even like this, and lixiviate is (seeing table 3) fully; 2. select for use the lixiviate fluid power of research screening to make to sea-tangle and do not cause interference between al species, Al (III) is extracted fully; 3., guard column CG5A 4 * 50mm of selecting for use, CG5A 4 * 250mm cation exchange column can make al species separate to see Fig. 2.Can be known by this spectrogram: 1. the form of sample Al after extract extracts does not change along with the increase of extraction time.2. the peak shape of AL (III) is highly stable, proves between al species can not transform each other with the increase of extraction time.

Embodiment 2: to the mensuration of Al in the sea cucumber (III):

Take by weighing bright appearance (need smash homogenate to pieces) 4.000g and place 50mL tool plug scale test tube, add moving phase 20mL mixing.To 70 ℃ of shaking bath 1h, make the abundant lixiviate of sample.Take out cooling.The centrifugal 15min of 4000r/min takes out supernatant with 0.45 micron water system filtering with microporous membrane, the value of last machine mensuration AL (III).Other condition is identical with sea-tangle all, the mensuration result of sea cucumber recovery of standard addition (seeing table 4).

The mensuration result of table 4, sea cucumber recovery of standard addition (in butt)

Can be known by table 4: we add the recovery of target Al (III) 90.1%, and recovering effect is desirable;

In sum, can be known by table 2: the analytical performance of this instrument meets the requirement of measuring AL (III) fully, can know the recovery that we add target Al (III) at 88.7%-90.1% by table 3 and table 4, the recovering effect ideal.This pre-treatment that has benefited from instrument that we select for use and instrument condition, extract concentration, sample very rationally puts in place.The bright spot of this method: the Al in aquatic products and the goods thereof (III) is after extract extracts; Separate through high performance liquid chromatograph; Because the cation-exchange chromatography post is different to the adsorptive power of the various forms of Al, moving phase is with its wash-out successively, and eluent is right through the inductively coupled plasma mass spectrometer ²⁷AL carries out analyzing and testing; This method detection line is low, sensitivity, accuracy height;

Therefore want to measure accurately the Al (III) in aquatic products and the goods thereof, must use high performance liquid chromatography-inductively coupled plasma mass spectrometer coupling (HPLC-ICP-MS) morphological analysis method.This method has provided the more complete biological characteristics of the aluminium in the aquatic products, and aluminium in the particularly current marine alga is measured and caused under the situation of dispute in many ways, and adopting more clear, the morphological analysis of HPLC-HGAFS coupling accurately is best first-selection.We are just presiding over the formulation of bearing China's agricultural industry criteria " assay method of trivalent aluminium and organo-aluminium in marine alga and the goods thereof " at present, and filling up with it does not still have this blank of detection method of measuring trivalent aluminium in aquatic products and the goods thereof in ISO and the CAC standard.

Claims (1)

1. the assay method of trivalent aluminium in aquatic products and the goods thereof; May further comprise the steps: instrument of 1, selecting for use and equipment, HPLC-ICP-MS combined system, 2, configuration reagent and standard solution; 3, the step of sample preparation, 4, the used instrument condition 5 of experiment, sample detection and result's calculating; It is characterized in that concrete steps are following:

1) instrument of selecting for use and equipment, the HPLC-ICP-MS combined system:

(1), HPLC part: use SERIES200 high-pressure liquid phase pump, be furnished with automatic sampler, be furnished with CG5A 4 * 250mm cation-exchange chromatography post of guard column CG5A 4 * 50mm and identical material filler;

(2), ICP-MS: be furnished with the dynamic response pond, sampling system, radio-frequency signal generator, plasma system, interface, ion lens system, quadrupole rod, vacuum system, detecting device, air-path control system;

(3), cooling circulating water;

(4), constant temperature water bath oscillator;

(5), sand core filter;

(6), circulating vacuum pump;

(7), hydro-extractor;

(8), numerical control ultrasonic cleaning machine;

(9), the water system miillpore filter of 0.45 μ m;

(10), the argon gas of purity >=99.99%;

2) configuration reagent and standard solution:

(1), leaching liquor: the mixed liquor of 0.0040mol/L lithium hydroxide, 0.0060mol/L pyridine-2.6-dicarboxylic acid, 0.090mol/L glacial acetic acid;

(2), moving phase: the mixed liquor of 0.0040mol/L lithium hydroxide, 0.0060mol/L pyridine-2.6-dicarboxylic acid, 0.090mol/L glacial acetic acid, before using through 0.45 μ m membrane filtration and ultrasonic degas;

All reagent all is that analysis is pure or top grade is pure, and ultrapure water is all used in whole experiment; Warp 15% nitric acid dousing 24h before glass apparatus uses;

(3), the standard of AL (III) is used liquid:

AL (III) standard is used liquid: the storing solution that 100 μ g/mL AL (III) standard solution is diluted to 1.00 μ g/mL AL (III); Putting 4 ℃ of refrigerators deposits subsequent use; AL (III) storing solution by 1.00 μ g/mL uses liquid through AL (III) standard that moving phase is diluted to 50.0ng/mL, 100.0ng/mL, 250.0ng/mL, 500.0ng/mL, 1000.0ng/mL, and is existing with join at present;

3) step of sample preparation:

Take by weighing through the dry sample 0.400g～0.800g that pulverized 40 mesh sieves and place 50mL tool plug scale test tube, add extract 20mL mixing, or bright appearance is smashed homogenate to pieces and taken by weighing 2.000g～4.000g and place 50mL tool plug scale test tube; Add extract 20mL mixing, to 70 ℃ of water bath with thermostatic control shaking table 1h, after the abundant lixiviate of sample; Take out cooling, the centrifugal 15min of 4000r/min takes out supernatant with 0.45 micron water system filtering with microporous membrane; Move in the sample introduction bottle, last machine is measured the value of trivalent aluminium;

4) the used instrument condition of experiment:

(1), high-efficient liquid phase chromatogram condition HPLC: moving phase is the mixed liquor of 0.0040mol/L lithium hydroxide, 0.0060mol/L pyridine-2.6-dicarboxylic acid, 0.090mol/L glacial acetic acid, flow velocity 1mL/min; Sampling volume 50～100 μ L, auto injection;

(2), inductively coupled plasma mass spectrometer condition ICP-MS: radio-frequency power 1150～1600W, plasma flow speed 15～20L/min, secondary air speed 1.2～1.5L/min; Sampling time 10min; Measure mass number ²⁷AL;

5) sample detection and result calculate:

(1), qualitative: the start back is provided with by above-mentioned condition, after waiting to stablize, does typical curve earlier, measures the sample measuring liquid of handling well of treating then; Under above-mentioned high performance liquid chromatography and inductively coupled plasma mass spectrometer condition, adopt full-automatic injector sample introduction 50～100 μ L to measure, high performance liquid chromatograph separates, and the inductively coupled plasma mass spectrometer is right ²⁷AL detects; AL under above-mentioned instrument condition (III) retention time is about 3.172min;

(2), quantitative: as to treat that the chromatographic peak retention time of AL in the sample measuring liquid (III) and standard use liquid phase within ± 10%, promptly to think this material to be determined than variation range;

The result calculates and statement:

With the external standard method in the data processing software, confirm AL (III) retention time and degree of separation pulse strength, be horizontal ordinate with concentration; The pulse strength integration is an ordinate; The drawing standard working curve then with appearance liquid peak analyzing and processing, can obtain treating the concentration of the AL (III) in the sample measuring liquid; Calculate the content of trivalent aluminium in the derived sample by following formula (1):

X＝C×V/m×1000............... (1)

In the formula:

The content of trivalent aluminium in the X-sample, mg/kg;

The concentration of trivalent aluminium in the C-liquid to be measured, ng/ml;

The m-sample weighting amount, g;

V-measures total liquid volume, mL.

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