US 11,747,349 B2
Method for simultaneously detecting vitamin K1 and vitamin K2 in traces of blood
Jinbao Zhao, Beijing (CN); Yongjuan Jia, Beijing (CN); and Junjun Ni, Beijing (CN)
Assigned to Beijing Harmony Health Medical Diagnostics Co., Ltd., Beijing (CN)
Appl. No. 17/257,088
Filed by Beijing Harmony Health Medical Diagnostics Co., Ltd., Beijing (CN)
PCT Filed Sep. 29, 2020, PCT No. PCT/CN2020/118947
§ 371(c)(1), (2) Date Dec. 30, 2020,
PCT Pub. No. WO2022/067533, PCT Pub. Date Apr. 7, 2022.
Prior Publication US 2022/0326261 A1, Oct. 13, 2022
Int. Cl. G01N 33/82 (2006.01); G01N 1/34 (2006.01); G01N 30/60 (2006.01); G01N 30/72 (2006.01); G01N 30/88 (2006.01); G01N 30/02 (2006.01)
CPC G01N 33/82 (2013.01) [G01N 1/34 (2013.01); G01N 30/6034 (2013.01); G01N 30/7233 (2013.01); G01N 30/88 (2013.01); G01N 2030/027 (2013.01); G01N 2030/8822 (2013.01)] 13 Claims
OG exemplary drawing
 
1. A method for simultaneously detecting Vitamin K1 and Vitamin K2 in blood, comprising:
(1) establishment of an analytical method for detecting Vitamin K1 and Vitamin K2 in blood, comprising selection of two-dimensional liquid chromatography-tandem mass spectrometer system modules, construction of a two-dimensional liquid chromatography-tandem mass spectrometer system, and establishment of analytical conditions of the two-dimensional liquid chromatography-tandem mass spectrometer,
(2) standardization of standard solutions
(2a) preparing at least three mixed standard solutions, wherein the mixed standard solution is a solution having an internal standard substance, Vitamin K1, and Vitamin K2, and the concentration of the internal standard substance in the at least three mixed standard solutions is the same;
(2b) detecting each of the at least three mixed standard solutions with the two-dimensional liquid chromatography-tandem mass spectrometer using the analytical method established in Step (1) to obtain first detection results respectively corresponding to the at least three mixed standard solutions; and
(2c) fitting standard curve equations respectively corresponding to Vitamin K1 and Vitamin K2 according to each of the first detection results and concentrations of the internal standard substance, Vitamin K1 and Vitamin K2 in the mixed standard solutions; and
(3) detection of a blood sample
(3a) adding the same amount of internal standard substance as in the mixed standard solution to the blood sample, adding an extraction reagent, performing centrifugation after the extraction, collecting a supernatant obtained after the centrifugation, blowing the supernatant to dry, and redissolving the residue with a redissolution solution to obtain an analytical sample;
(3b) detecting the blood sample with the two-dimensional liquid chromatography-tandem mass spectrometer using the analytical method established in Step (1) to obtain a second detection result corresponding to the blood sample; and
(3c) obtaining concentrations of Vitamin K1 and Vitamin K2 in the blood sample based on the second detection result and the standard curve equations respectively corresponding to Vitamin K1 and Vitamin K2;
wherein the establishment of the analytical conditions of the two-dimensional liquid chromatography-tandem mass spectrometer in Step (1) comprises:
selecting phenylhexyl as the first dimensional chromatographic column of two-dimensional liquid chromatography; and C18 as the second dimensional chromatographic column.