Method for detecting multiple lipid-soluble vitamins in blood sample simultaneously
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A fat-soluble vitamin, blood sample technology, applied in measuring devices, instruments, scientific instruments, etc., can solve the problems of low reliability of clinical testing, affecting the accuracy of test results, and the amount of blood required for testing, so as to reduce blindness. Vitamin supplementation, objective and easy results, reliable results
Active Publication Date: 2015-12-16
JINAN YING SHENG BIOTECH
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The antibodies used in immunoassay detection can also recognize other vitamin derivatives, and the antibody-antigen reaction is usually less than 100%, so the results vary greatly and the specificity is poor, and the reliability of clinical detection is not high, especially for the detection of content lower vitamin
The current liquid chromatography-mass spectrometry can only do qualitative analysis on one or two vitamins. Due to the complex composition of plasma/serum and the low content of vitamin...
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Embodiment 1
[0045] (1) Prepare the sample to be tested
[0046] Add absolute ethanol to the serum sample to precipitate the protein, then add n-hexane to extract, vortex and centrifuge, take the n-hexane layer into the sample bottle, dry it with nitrogen, and redissolve the sample in 70% v / v methanol aqueous solution. Filter through a 0.2 μm filter membrane to obtain the sample to be tested.
[0047] Wherein, the volume ratio of serum, absolute ethanol, and n-hexane is 1:2:5, and the volume ratio of serum to reconstituted solution is 2:1.
[0049] The sample to be tested enters the chromatographic column for separation through the gradient elution mode, and the reference conditions of liquid chromatography are:
[0077] Serum samples were obtained from patients diagnosed with vitamin D deficiency in the hospital.
[0078] The specific steps are the same as in Example 1, and the volume ratio of serum, absolute ethanol, and n-hexane is 1:2:5.
[0079] By calculation, the content of vitamin 25(OH)D2 and 25(OH)D3 in patients was significantly lower than that in healthy people.
[0080] Experiments have proved that the invention has simple sample processing, high throughput and low cost, and can simultaneously detect six kinds of fat-soluble vitamins in the human body, which has very important significance.
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Abstract
The invention discloses a method for detecting multiple lipid-soluble vitamins in a blood sample simultaneously. The method comprises the steps of preprocessing a biological sample by a simple liquid-liquid extraction method, then carrying out chromatographic separation and mass spectrometric detection, respectively selecting a pair of qualitative ions and a pair of quantitative ions according to each vitamin, wherein the relative retention time and qualitative ion pairs of the various vitamins are taken as qualitative basis, and a standard curve for quantitation is formed by standard substances. Meanwhile, according to the method, three levels of quality control substances are used for inspecting the accuracy and effectiveness of the method, and the distortion of the detection result is avoided. According to the method, the purpose that six lipid-soluble vitamins in one serum sample can be simultaneously detected can be realized for the first time by applying an LC-MS technology, the influence caused by interferents can be alleviated, the operation is simple, convenient and rapid, the analysis time is only 4.8min, the throughput is high, the cost is low, the levels of lipid-soluble vitamins in a human body are effectively monitored, and the method has guiding significance on the reasonable and safe supplementation of vitamins, and is liable to clinical promotion and popularization.
Description
technical field [0001] The invention belongs to the field of vitamin detection, in particular to a method for simultaneously detecting multiple fat-soluble vitamins in blood samples. Background technique [0002] Vitamins are micronutrients necessary for biological growth and metabolism. They are divided into fat-soluble and water-soluble vitamins. When they are deficient, organisms cannot grow normally and lead to vitamin deficiency. Vitamin A, 25(OH)D2, 25(OH)D3, E, K1, and K2 are all fat-soluble vitamins. When the human body lacks these six vitamins, it will show different symptoms: vitamin A deficiency can lead to night blindness, eye inflammation Etc.; vitamin D deficiency can lead to rickets in children, adult osteomalacia, osteoporosis in the elderly, etc.; vitamin E deficiency can cause neonatal hemolytic anemia, nervous system diseases, etc.; vitamin K deficiency can cause coagulation dysfunction, blood coagulation Prolonged time and bleeding etc. [0003] In fore...
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