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Method for detecting content of methylmalonic acid in blood

A technology of methylmalonic acid and blood, applied in the field of clinical chemistry, can solve the problems of cumbersome derivatization treatment operations, expensive ultrafiltration centrifuge tube consumables, and high detection costs

Pending Publication Date: 2021-05-25
BEIJING HARMONY HEALTH MEDICAL DIAGNOSTICS CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the operation of derivatization treatment is relatively cumbersome, and the operation of ultrafiltration centrifugation pretreatment is relatively simple, but the consumables of ultrafiltration centrifuge tubes are expensive, resulting in high detection costs

Method used

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  • Method for detecting content of methylmalonic acid in blood
  • Method for detecting content of methylmalonic acid in blood
  • Method for detecting content of methylmalonic acid in blood

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0086] The embodiment of the present invention is used to obtain the standard curve equation.

[0087] 1.1 Preparation of standard stock solution

[0088] Purchasing the standard solution of methylmalonic acid, that is, dissolving the standard methylmalonic acid in acetonitrile to obtain a standard stock solution, wherein the concentration of the standard methylmalonic acid is 1mg / mL and stored at -80°C Stored under certain conditions, the validity period is 1 year.

[0089] 1.2 Preparation of internal standard stock solution

[0090] Accurately weigh 2mg of methylmalonic acid-d3 standard substance in a 2mL volumetric flask, add an aqueous solution containing 50% methanol to dissolve and set the volume to 2mL to obtain an internal standard stock solution, wherein, methylmalonic acid-d3 standard substance The concentration is 1000μg / mL, and stored at -80°C, the validity period is 1 year.

[0091] 1.3 Instruments for testing

[0092] High performance liquid chromatography ta...

Embodiment 2

[0126] The embodiment of the present invention is used for detecting the content of methylmalonic acid in blood.

[0127] 2.1 Obtaining blood samples

[0128] Take at least 1 mL of the blood to be tested, centrifuge at a centrifugation speed of 3500 rpm for 10 min, and take the supernatant to obtain serum, which is used as a blood sample and stored at 4°C until analysis.

[0129] After the blood sample is obtained, it can be pre-treated to obtain a corresponding sample to be tested that can be directly loaded.

[0130] 2.2 Blood sample pretreatment

[0131]Use a pipette gun to pipette 100 μL of serum into a 1.5 mL centrifuge tube, then add 10 μL of internal standard working solution, vortex and mix at 2500 rpm for 1 min; then add 300 μL of acetonitrile, and vortex and mix at 2500 rpm for 3 min, Centrifuge at a high speed of 12000rpm for 5min, pipette 350μL of the supernatant to another 1.5mL centrifuge tube; blow dry the supernatant with nitrogen at 55℃, then redissolve with...

Embodiment 3

[0142] The embodiment of the present invention is used to detect the content of methylmalonic acid in blood plasma.

[0143] In the above step 2.1, the blood to be tested is processed to obtain serum. Different from this, in the embodiment of the present invention, the blood to be tested is processed to obtain plasma, and the obtained plasma is used as a blood sample, while other processing procedures are consistent with the above-mentioned embodiment 2.

[0144] By detecting the content of methylmalonic acid in plasma, it was found that the obtained detection results were consistent with the detection results obtained in Example 2 within the allowable error range.

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Abstract

The invention provides a method for detecting the content of methylmalonic acid in blood, wherein the comprises the following steps: under certain detection conditions, respectively detecting at least three standard solutions by using a high performance liquid tandem mass spectrometer to obtain chromatograms of the standard solutions, wherein each standard solution contains a methylmalonic acid standard substance with known concentration and an internal standard substance; fitting according to the chromatograms of the standard solutions to obtain a standard curve equation; uniformly mixing the internal standard working solution and a blood sample, adding acetonitrile, uniformly mixing, centrifuging to obtain a supernatant, blow-drying the supernatant in a heating and nitrogen blowing manner, adding an aqueous solution containing 0-2% of formic acid and 0-20% of acetonitrile after blow-drying, uniformly mixing, and centrifuging to obtain a supernatant to obtain a to-be-detected sample; detecting the to-be-detected sample to obtain a chromatogram of the to-be-detected sample; and calculating the content of methylmalonic acid in the blood sample according to the chromatogram of the to-be-detected sample and the standard curve equation. According to the scheme, the method for detecting the content of methylmalonic acid in blood is provided, the sample pretreatment operation is simple, and the cost is low.

Description

technical field [0001] The invention relates to the technical field of clinical chemistry, in particular to a method for detecting the content of methylmalonic acid in blood. Background technique [0002] According to epidemiological survey data, the vitamin B12 deficiency rate is about 20% among surveyed populations in the United States and Canada, up to 40% in Latin American countries, and even as high as 80% among Indian children. Women of childbearing age, infants and the elderly are high-risk groups for vitamin B12 deficiency. Vitamin B12 deficiency can cause blood system, metabolism and nervous system lesions, especially increase the birth risk of pregnant women, and affect the cognitive development of infants and young children. Studies have confirmed that some elderly people with mildly reduced serum vitamin B12 have methylmalonic acid (MMA, molecular formula C 4 h 6 o 4 ) has been significantly increased, suggesting that the determination of metabolite MMA is mo...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N30/88
CPCG01N30/88G01N2030/8822G01N2030/045G01N2030/027Y02A50/30
Inventor 邱天祎贾永娟许丽倪君君
Owner BEIJING HARMONY HEALTH MEDICAL DIAGNOSTICS CO LTD
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