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Derivatization-based detection method for 12 steroid hormones in dried blood spot

A technology for steroid hormones and detection methods, which is applied in the detection field of 12 steroid hormones in dried blood spots, can solve the problems that the lower detection limit cannot meet the clinical needs and the ionization efficiency is not high, and can improve the ionization process, improve the detection sensitivity, reduce the invasive effect

Pending Publication Date: 2022-07-05
天津国科医疗科技发展有限公司
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Problems solved by technology

However, the dried blood spot technique has rarely been reported in the sample analysis of endogenous steroid hormones
Patent CN 107462653A discloses a liquid chromatography tandem mass spectrometry detection method for 7 steroid hormones in dried blood spots, which can be analyzed after extracting the dried blood slices with methanol, but the ionization efficiency of the indicators is not high, and the lower limit of detection cannot be satisfied clinical needs

Method used

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  • Derivatization-based detection method for 12 steroid hormones in dried blood spot
  • Derivatization-based detection method for 12 steroid hormones in dried blood spot
  • Derivatization-based detection method for 12 steroid hormones in dried blood spot

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Experimental program
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Effect test

Embodiment 1

[0056] 1. Reagent source description

[0057] Standard: Dihydrotestosterone (DHT), Dehydroepiandrosterone (DHEA), 17-hydroxyprogesterone (17-OHP), Progesterone (P), Corticosterone (B), Cortisone (E), Aldosterone (Ald), estrone (E1), and estradiol (E2) were purchased from Sigma-Aloric Company, and estriol (E3) and 17-hydroxypregnenolone (17-OHPreg) were purchased from Shanghai Zhenzhun Biotechnology Co., Ltd. Company, androstenedione (AD) was purchased from Beijing Zhenxiang Technology Co., Ltd.; progesterone- 13 C 3 Purchased from Sigma-Aloric, androstenedione- 13 C 3 , 17-hydroxypregnenolone-d 2 13 C 2 Purchased from Shanghai Zhenzhun Biotechnology Co., Ltd., DHT- 13 C 3 , DHEA- 13 C 3 , 17-hydroxyprogesterone- 13 C 3 , corticosterone-d 8 , Cortisone- 13 C 3 , estrone- 13 C 3 , estradiol-d 4 , estriol-d 3 , aldosterone-d 8 It was purchased from Shanghai Pufen Biotechnology Co., Ltd. Methanol and acetonitrile were purchased from Tianjin Concord Company. A...

specific Embodiment approach

[0110] In the experiment, the peak area response of the index after the same sample was processed by the dried blood spot technology and the conventional liquid-liquid extraction was also explored. Specific implementation: take a whole blood sample and divide it into two parts, one of which is subjected to extraction and derivatization according to the dry blood spot technology and then tested on the machine; the other is taken after low-temperature centrifugation to take 45 μL of upper plasma for liquid-liquid extraction, and 1 mL of formazan is added. After the solvent was completely removed, 80 μL of methanol-water (1 / 1, v / v) solution was added, and the solution was thoroughly mixed and dissolved, and then tested on the machine (liquid-liquid). The extraction was the same quantitation ion pair used for the underivatized carbonyl steroid in (4). The peak area data of the comparative steroid indicators are shown in Table 5:

[0111] table 5

[0112] Steroid drie...

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Abstract

The invention discloses a derivatization-based method for detecting 12 steroid hormones in dried blood spots, which comprises the following steps: 1) pretreatment of a dried blood spot sample: 1-1) extraction: taking a blood spot from the dried blood spot sample, adding an internal standard extract liquor, and oscillating; 1-2) nitrogen blowing: centrifuging and then nitrogen blowing to dry; (1-3) deriving: adding an acetonitrile aqueous solution containing hydroxylamine hydrochloride, uniformly mixing, and deriving at constant temperature to prepare a test sample; and 2) detecting the test sample by adopting a liquid chromatography-tandem mass spectrometry method, and calculating the contents of the 12 steroid hormones in the dried blood spot sample to be detected through a standard curve. The adopted dried blood spot sampling has extremely small invasiveness, the sample collection is simple and convenient, the matrix effect of the biological sample can be effectively removed, and the dried blood spots are convenient to transport and store; according to the method, the ionization efficiency of carbonyl steroid hormone can be improved through hydroxylamine derivation, and the ionization process of hydroxyl steroid can be improved by adding ammonium fluoride into the mobile phase.

Description

technical field [0001] The invention relates to the technical field of clinical mass spectrometry in vitro detection, in particular to a method for detecting 12 steroid hormones in derivatized dried blood spots. Background technique [0002] Steroid hormones include glucocorticoids, mineralocorticoids and sex hormones. Their physiological functions are mainly manifested in regulating glucose metabolism, regulating water and salt metabolism, and regulating gonadal function. clear role. Steroid analysis by LC-MS / MS has been widely used in the screening of congenital adrenal hyperplasia, primary aldosteronism, Cushing's syndrome, polycystic ovary syndrome, and other diseases. [0003] At present, human serum or plasma samples are processed for endogenous steroid hormone analysis by liquid-liquid extraction, protein precipitation, solid-phase extraction, etc., but invasive venous blood collection and sample collection are required for serum or plasma steroid analysis. , the pr...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N30/02G01N30/06G01N30/34G01N30/60G01N30/72G01N30/86
CPCG01N30/02G01N30/06G01N30/34G01N30/6052G01N30/7266G01N30/86G01N2030/047G01N2030/062G01N2030/067G01N2030/045
Inventor 李艳杰覃素姿周传贵王天一李艳胡玮程文播
Owner 天津国科医疗科技发展有限公司
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