CN103869024B - Children's's calcium carbonate D3 particle vitamin d3 levels detection method - Google Patents
Children's's calcium carbonate D3 particle vitamin d3 levels detection method Download PDFInfo
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Abstract
The invention discloses a kind of detection method being applicable to vitamine D3 in children's's calcium carbonate D3 particle.Comprise: comprise the steps: that (1) precision takes children's's calcium carbonate D3 particle, put in centrifuge tube, make the solution containing BHT 0.2mg in every 1ml with dimethyl sulfoxide (DMSO)-water (10 ﹕ 2) mixed solution; (2) by ultrasonic for step (1) gained solution, the isooctane solution of BHT is added, jolting, centrifugal, divide and get upper solution, as test sample; (3) vitamine D3 reference substance is separately got appropriate, accurately weighed, dissolve with isooctane solution and make the solution about containing 1 μ g in every 1ml, being measured in the same method; (4) by external standard method with calculated by peak area, result of calculation is multiplied by 1.09 and namely obtains vitamin d3 levels.Operability is stronger, accuracy is higher.
Description
Technical field
The present invention relates to the detection method that a kind of medicine is called vitamine D3 in children's's calcium carbonate D3 particle, belong to Pharmaceutical Analysis technical field.
Background technology
Children's's calcium carbonate D3 particle is that a kind of children replenish the calcium medicine, every bag containing 0.75 gram, calcium carbonate (being equivalent to calcium 0.3 gram), vitamine D3 100 international unit (2.5 microgram); Researched and developed by ounce pharmaceutical Co. Ltd of the U.S. the earliest, produced listing in 2009, and sell to domestic packing in import in the same year, trade name " D-Cal ", operative norm is import standard (standard No. is JX20070088).
Detection method in import standard under vitamin d3 levels mensuration item is as follows:
Chromatographic condition and system suitability: be filling agent with amino bonded silica gel; With normal hexane-isopropyl alcohol (99 ﹕ 1) for mobile phase, flow velocity is 1.5ml per minute, and determined wavelength is 265nm.Get vitamine D3 reference substance solution to heat in 60 DEG C of water-baths 1 hour (making generation PreD3), let cool, measure 20 μ l, injection liquid chromatography, record chromatogram, the degree of separation at vitamine D3 peak and PreD3 peak all should be greater than 5, and number of theoretical plate calculates should be not less than 5000 by vitamine D3 peak, and tailing factor should be not more than 2.0.
Determination method: get children's's calcium carbonate D3 particle 25 bags of contents, accurately weighed, calculate every bag of average loading amount, porphyrize, precision takes fine powder appropriate (being about equivalent to vitamine D3 25 μ g), put in the brown measuring bottle of 100ml, add solution [to make in every 1ml containing 2 with dimethyl sulfoxide (DMSO)-water (10 ﹕ 2) mixed solution, the solution of 6-BHT 0.2mg] 50ml, ultrasonic 15 minutes, ultrasonic water bath temperature is kept to be no more than 30 DEG C, precision adds 0.02%2, the hexane solution 25ml of 6-BHT, jolting is after 15 minutes, centrifugal 5 minutes (4000rpm), divide and get upper solution, lucifuge leaves standstill after 1 hour and obtains settled solution as need testing solution, precision measures 100 μ l, injection liquid chromatography, record chromatogram, separately get vitamine D3 reference substance appropriate, accurately weighed, with 0.02%2, the hexane solution of 6-BHT dissolves and makes the solution about containing 1 μ g in every 1ml, be measured in the same method, by external standard method with calculated by peak area, result of calculation is multiplied by 1.09 and namely obtains vitamin d3 levels.
Adopt during the determination method processing sample solution of above-mentioned detection method and run into following problem:
1. the centrifuge tube volume that hydro-extractor is supporting is not enough;
2. sample solution can not shift completely when brown measuring bottle is transferred to centrifuge tube, and residue is more;
3. normal hexane boiling point is only 68.7 DEG C, easily volatilizees when room temperature is higher, there is larger environmental pollution and safety problem;
4. after centrifugal, can not also need standing 1 hour by sample introduction immediately.
Summary of the invention
The object of this invention is to provide the determination method that a kind of operability is stronger, accuracy is higher, for detecting the vitamin d3 levels in children's's calcium carbonate D3 particle.
Inventor improves as follows to existing determination method: 1. carry out adjustment sample volume and solvent load according to physical condition; 2. brown measuring bottle changes centrifuge tube (centrifuge tube material is teflon or other inertia materials) into; 3. normal hexane changes isooctane (boils up till 99.3 DEG C) into; 4. extend centrifugation time, without the need to leaving standstill after centrifugal.
The present invention is as follows to the determination method of vitamin d3 levels in children's's calcium carbonate D3 particle: (1) precision takes children's's calcium carbonate D3 particle, put in centrifuge tube, the solution containing BHT 0.2mg in every 1ml is made with dimethyl sulfoxide (DMSO)-water (10 ﹕ 2) mixed solution; (2) by the ultrasonic 15min of step (1) gained solution, the isooctane solution of 0.02%2,6-BHT is added, jolting, centrifugal, divide and get upper solution, as test sample, precision measures 100 μ l, injection liquid chromatography, record chromatogram; (3) vitamine D3 reference substance is separately got appropriate, accurately weighed, dissolve with the isooctane solution of 0.02%2,6-BHT and make the solution about containing 1 μ g in every 1ml, being measured in the same method; (4) by external standard method with calculated by peak area, result of calculation is multiplied by 1.09 and namely obtains vitamin d3 levels.
Concrete implementation step: get children's's calcium carbonate D3 particle 20 bags of contents, accurately weighed, calculate every bag of average loading amount, porphyrize, precision takes fine powder appropriate (being about equivalent to vitamine D3 10 μ g), put in 50ml centrifuge tube (centrifuge tube material is teflon or other inertia materials), add solution [to make in every 1ml containing 2 with dimethyl sulfoxide (DMSO)-water (10 ﹕ 2) mixed solution, the solution of 6-BHT 0.2mg] 20ml, ultrasonic 15 minutes, ultrasonic water bath temperature is kept to be no more than 30 DEG C, precision adds 0.02%2, the isooctane solution 10ml of 6-BHT, jolting is after 15 minutes, centrifugal 10 minutes (4000rpm), upper solution is got without the need to leaving standstill directly to divide, as need testing solution, precision measures 100 μ l, injection liquid chromatography, record chromatogram, separately get vitamine D3 reference substance appropriate, accurately weighed, with 0.02%2, the isooctane solution of 6-BHT dissolves and makes the solution about containing 1 μ g in every 1ml, be measured in the same method, by external standard method with calculated by peak area, result of calculation is multiplied by 1.09 and namely obtains vitamin d3 levels.
1. chromatographic condition and system suitability: be filling agent with amino bonded silica gel; With normal hexane-isopropyl alcohol (99 ﹕ 1) for mobile phase, flow velocity is 1.5ml per minute, and determined wavelength is 265nm.Get vitamine D3 reference substance solution to heat in 60 DEG C of water-baths 1 hour (making generation PreD3), let cool, measure 20 μ l, injection liquid chromatography, record chromatogram, the degree of separation at vitamine D3 peak and PreD3 peak all should be greater than 5, and number of theoretical plate calculates should be not less than 5000 by vitamine D3 peak, and tailing factor should be not more than 2.0.
2. the preparation of need testing solution: get children's's calcium carbonate D3 particle 20 bags of contents, accurately weighed, calculate every bag of average loading amount, porphyrize, precision takes fine powder appropriate (being about equivalent to vitamine D3 10 μ g), put in 50ml centrifuge tube (centrifuge tube material is teflon or other inertia materials), add solution [to make in every 1ml containing 2 with dimethyl sulfoxide (DMSO)-water (10 ﹕ 2) mixed solution, the solution of 6-BHT 0.2mg] 20ml, ultrasonic 15 minutes, ultrasonic water bath temperature is kept to be no more than 30 DEG C, precision adds 0.02%2, the isooctane solution 10ml of 6-BHT, jolting is after 15 minutes, centrifugal 10 minutes (4000rpm), upper solution is got without the need to leaving standstill directly to divide.
3. reference substance solution: get vitamine D3 reference substance appropriate, accurately weighed, dissolve with the isooctane solution of 0.02%2,6-BHT and make the solution about containing 1 μ g in every 1ml.
4. measure: be filling agent with amino bonded silica gel; With normal hexane-isopropyl alcohol (99 ﹕ 1) for mobile phase, flow velocity is 1.5ml per minute, and determined wavelength is 265nm.Precision measures above-mentioned reference substance solution and each 100 μ l of need testing solution respectively, injection liquid chromatography, record chromatogram; By external standard method with calculated by peak area, result of calculation is multiplied by 1.09 and namely obtains vitamin d3 levels.
Embodiment
The present invention is illustrated further below by a concrete example.
It is as follows that sample preparation and detection operate concrete steps:
1. the preparation of mobile phase: get chromatographically pure normal hexane 1000ml, isopropyl alcohol 500ml filters with miillpore filter respectively.
2. the preparation of solvent:
Dimethyl sulfoxide (DMSO)-water mixed solution: get chromatographically pure dimethyl sulfoxide (DMSO) 500ml, purified water 100ml, put in same beaker and mix, adds analytically pure BHT 120mg and makes dissolving.
The isooctane solution of 0.02%2,6-BHT is molten: get chromatographically pure isooctane 100ml, adds analytically pure BHT 20mg and makes dissolving.
3. the preparation of reference substance solution: get vitamine D3 reference substance 10mg, accurately weighed, put in 100ml measuring bottle, dissolve with the isooctane solution of 0.02%2,6-BHT; Precision measures 1ml again, puts in 100ml measuring bottle, is diluted to scale with the isooctane solution of 0.02%2,6-BHT, makes the solution about containing 1 μ g in every 1ml.
4. the preparation of system suitability solution: the reference substance solution that takes a morsel heats 1 hour (making generation PreD3) in 60 DEG C of water-baths, lets cool.
5. the preparation of need testing solution: get children's's calcium carbonate D3 particle 20 bags of contents, accurately weighed, calculate every bag of average loading amount, porphyrize, precision takes fine powder 4g (being about equivalent to vitamine D3 10 μ g), put in 50ml centrifuge tube, add dimethyl sulfoxide (DMSO)-water mixed solution 20ml, ultrasonic 15 minutes, keep ultrasonic water bath temperature to be no more than 30 DEG C, precision added 0.02%2, the isooctane solution 10ml of 6-BHT, jolting is after 15 minutes, and centrifugal 10 minutes (4000rpm), gets upper solution without the need to leaving standstill directly to divide.
6. system suitability: be filling agent with amino bonded silica gel; With normal hexane-isopropyl alcohol (99 ﹕ 1) for mobile phase, flow velocity is 1.5ml per minute, and determined wavelength is 265nm.Measure system suitability solution 20 μ l, injection liquid chromatography, record chromatogram, the degree of separation at vitamine D3 peak and PreD3 peak all should be greater than 5, and number of theoretical plate calculates should be not less than 5000 by vitamine D3 peak, and tailing factor should be not more than 2.0.
7. the mensuration of vitamin d3 levels in test sample: precision measures above-mentioned reference substance solution and each 100 μ l of need testing solution respectively, injection liquid chromatography, record chromatogram; By external standard method with calculated by peak area, result of calculation is multiplied by 1.09 and namely obtains vitamin d3 levels.Computing formula is as follows:
A in formula
right: vitamin D in reference substance solution chromatogram
3peak area;
A
supply: vitamin D in need testing solution chromatogram
3peak area;
C
right: vitamin D in reference substance solution
3concentration (μ g/ml);
M
all: children's's calcium carbonate D
3the average loading amount (g/ bag) of particle;
M
supply: children's's calcium carbonate D
3particle sampling amount (g);
The isooctane solution volume (ml) of 5:0.02%2,6-BHT;
2.5: the vitamin D that every 1 bag (1g/ bag) contains
3amount (μ g);
1.09: vitamin D
3with provitamin D
3be converted to by vitamin D
3the correction factor of meter.
8. replica test Comparative result
Adopt method and the inventive method in aforesaid standards to carry out replica test respectively, carry out assay with legal system for 6 increment product, carry out the contrast of precision.Result is as following table:
Result shows, the result relative standard deviation (RSD) that the inventive method records is less, and precision is better.
9. recovery test Comparative result
Adopt method and the inventive method in aforesaid standards to carry out recovery test respectively, prepare each 3 increment product respectively at 80%, 100%, 120% concentration level and carry out assay, carry out the contrast of accuracy.Result is as following table:
Result shows, the recovery recorded according to the inventive method is closer to 100%, and relative standard deviation (RSD) is less, and accuracy is better.
Claims (1)
1. the detection method of vitamine D3 in children's's calcium carbonate D3 particle, is characterized in that comprising the steps:
(1) precision takes children's's calcium carbonate D3 particle, puts in centrifuge tube, adds dimethyl sulfoxide (DMSO): the mixed solution of water=10 ﹕ 2, containing BHT 0.2mg in mixed solution described in every 1ml;
(2) by the ultrasonic 15min of step (1) gained solution, the isooctane solution of 0.02%2,6-BHT is added, jolting, centrifugal, divide and get upper solution, as test sample, precision measures 100 μ l, injection liquid chromatography, record chromatogram;
(3) vitamine D3 reference substance is separately got appropriate, accurately weighed, dissolve with the isooctane solution of 0.02%2,6-BHT and make the solution about containing 1 μ g in every 1ml, being measured in the same method;
(4) by external standard method with calculated by peak area, result of calculation is multiplied by 1.09 and namely obtains vitamin d3 levels.
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| CN106290619A (en) * | 2016-07-31 | 2017-01-04 | 合肥远志医药科技开发有限公司 | A kind of child ties up the detection method of vitamin d3 levels in D Biocal preparation |
| CN105974035A (en) * | 2016-07-31 | 2016-09-28 | 合肥远志医药科技开发有限公司 | Method for detecting content of vitamin D3 in vitamin D calcium chewable tablet preparation |
| CN109490435A (en) * | 2018-11-02 | 2019-03-19 | 济南维瑞医药科技开发有限公司 | A kind of method of vitamine D3 impurity in detection chewing compound calcium carbonate tablet |
| CN109541110A (en) * | 2018-12-07 | 2019-03-29 | 浙江核力欣健药业有限公司 | A kind of detection children's calcium carbonate D3Vitamin D in particle3Method in relation to substance |
| CN109709237A (en) * | 2018-12-07 | 2019-05-03 | 浙江核力欣健药业有限公司 | A kind of calcium carbonate D3The related substance detecting method of chewable tablets |
| CN109813592A (en) * | 2019-03-26 | 2019-05-28 | 济南维瑞医药科技开发有限公司 | The identification pre-treating method of vitamine D3 in one vegetable oil |
| CN114088826A (en) * | 2021-10-27 | 2022-02-25 | 昆明源瑞制药有限公司 | Extraction method of vitamin D3 and method for detecting content of vitamin D3 |
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Denomination of invention: Method for detecting content of vitamin D3 in infant calcium carbonate D3 granule Effective date of registration: 20160927 Granted publication date: 20160120 Pledgee: Bank of Guiyang Limited by Share Ltd. Zunyi branch Pledgor: Guizhou Liansheng Pharmaceutical Co.,Ltd. Registration number: 2016990000821 |
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Denomination of invention: Method for detecting vitamin D3 content in pediatric calcium carbonate D3 granules Effective date of registration: 20230605 Granted publication date: 20160120 Pledgee: Bank of Guiyang Limited by Share Ltd. Zunyi branch Pledgor: Guizhou Liansheng Pharmaceutical Co.,Ltd. Registration number: Y2023980042754 |