Method for detecting vanillylmandelic acid in human urine
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A technology of vanillylmandelic acid and human urine, which is applied in the field of medical testing, can solve the problems of high price, high cost, and clinical difficulty, and achieve the effect of simple operation and low cost
Inactive Publication Date: 2015-07-29
HANGZHOU NIUBEI BIOTECH
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These methods have the advantages of high sensitivity and fast detection speed, but require special instruments, are expensive, costly, operators need special training, and are difficult to b...
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[0039] 1) Preparation of crude extract of VMA
[0040] a) After collecting fresh urine, add 5M HCl to acidify it, and store it in a -80°C freezer until subsequent experiments begin;
[0041] b) After the urine has melted, take 4ml of urine into a 15ml centrifuge tube pre-filled with 2g NaCl, mix upside down for 10min to form a saturated saline solution;
[0042] c) Add 4ml of ethyl acetate, mix upside down for 5min; centrifuge at 4000g for 5min, carefully absorb the upper organic phase into a new 50ml round bottom centrifuge tube;
[0043] d) Repeat steps b-c twice to obtain a total of 12ml of the upper organic phase;
[0044] e) Heat the centrifuge tube to 45°C with a metal heater, and volatilize under high wind in the fume hood until all the ethyl acetate is completely volatilized, and a yellow or black precipitate is obtained;
[0045] f) Add 400 μl of 1% acetic acid solution, shake fully, and dissolve overnight at 4°C;
[0046] g) After centrifugation at 15000g, transfe...
Embodiment 1
[0068] Embodiment 1: VMA detects figure such as image 3 shown.
[0069] Ret. Time
Conc.
area
Height
11.429
1.14448
238426
17914
[0070] The calculated content of VMA per 100 μl of the sample on HPLC was 0.414 μg.
[0071] The VMA content in urine was calculated to be 0.414μg / ml.
Embodiment 2
[0072] Embodiment 2: VMA detects figure such as Figure 4 Shown:
[0073] Ret. Time
Conc.
area
Height
11.414
2.15421
458154
32338
[0074] The calculated content of VMA per 100 μl of the sample on HPLC was 0.659 μg.
[0075] The VMA content in urine was calculated to be 0.659μg / ml.
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Abstract
The present invention relates to a method for detecting vanillylmandelic acid in human urine. The method comprises: taking human urine, and carrying out acidification corrosion protection with HCl so as to long-timely store at a temperature of -80 DEG C; extracting the urine with ethyl acetate, volatilizing, dissolving with acetic acid, and carrying out other processes so as to obtain a VMA crude extract; and separating the crude extract through HPLC, obtaining the stable and single VMA absorption peak through a UV detector at a wavelength of 280 nm, drawing a VMA standard curve through an external standard method, and finally calculating the VMA content. The method of the present invention has characteristics of low cost and easy operation, and provides a rapid, inexpensive and sensitive method for detecting VMA in human urine.
Description
technical field [0001] The invention relates to the field of medical detection, in particular to a method for detecting vanillylmandelic acid in human urine. Background technique [0002] Catecholamines are a group of hormones secreted by the adrenal medulla, including dopamine, epinephrine, and norepinephrine. These hormones, often in response to physical or mental stress, are released into the blood circulation, enhance the transmission of nerve impulses in the brain, release glucose and fatty acids for energy, bronchodilation and pupil dilation. Most catecholamines will be metabolized and degraded into vanilmandelic acid (VMA), homovanillic acid (HVA), metanephrine and norepinephrine, and only a small amount of catecholamines are excreted in urine . [0003] VMA, also known as 3-methoxy-4-hydroxymandelic acid or 4-hydroxy-3-methoxymandelic acid, is a metabolite of catecholamines epinephrine and norepinephrine, and is finally excreted in urine. The content of VMA in the...
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