Liquid Chromatography-Mass Spectrometry Methods For Multiplexed Detection and Quantitation of Free Amino Acids
AssigneeThe Boards of Trustees of the Leland Stanford Junior University
InventorsAnthony Le, Tina Cowan
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Abstract
The present invention provides liquid chromatography-mass spectrometry methods and kits for the specific, sensitive and rapid detection as well as quantitation of free amino acids in samples following High Pressure Liquid Chromatography (HPLC) separation.
Claims27
A method of detecting the presence of one or more free amino acids in a sample using liquid chromatography-mass spectrometry, comprising
- deproteinizing said sample to obtain free amino acids;
- separating the one or more free amino acids by liquid chromatography using strong ion-pairing conditions followed by weak ion-pairing conditions;
- ionizing said sample to generate ions;
- fragmenting ions to produce parent and daughter ions;
- detecting parent and daughter ions by selected reaction monitoring (SRM);
- and
- detecting the presence of one or more free amino acids in accordance to the detected parent and/or daughter ions.
A method of quantifying one or more free amino acids in a sample using liquid chromatography-mass spectrometry, comprising
- deproteinizing said sample to obtain free amino acids;
- separating the one or more free amino acids by liquid chromatography using strong ion-pairing conditions followed by weak ion-pairing conditions;
- ionizing said sample to generate ions;
- fragmenting ions to produce parent and daughter ions;
- detecting parent and daughter ions by selected reaction monitoring (SRM);
- and
- quantifying the one or more free amino acids in accordance to the detected parent and/or daughter ions.
A method of diagnosing a metabolic disorder in a subject, characterized by elevated or decreased levels of one or more amino acids, the method comprising the detection and quantitation of one or more free amino acids in a sample from said subject:
- comprising
- deproteinizing said sample to obtain free amino acids;
- separating the one or more free amino acids by liquid chromatography using strong ion-pairing conditions followed by weak ion-pairing conditions;
- ionizing said sample to generate ions;
- fragmenting ions to produce parent and daughter ions;
- detecting parent and daughter ions by selected reaction monitoring (SRM);
- and
- quantifying the concentration of said one or more free amino acids in accordance to the detected parent and/or daughter ions.
A method of monitoring a subject's response to the treatment of a metabolic disorder, characterized by normalized levels of one or more free amino acids in samples obtained some time after treatment of said subject, the method comprising
- deproteinizing said samples to obtain free amino acids;
- separating the one or more free amino acids by liquid chromatography using strong ion-pairing conditions followed by weak ion-pairing conditions;
- ionizing said samples to generate ions;
- fragmenting ions to produce parent and daughter ions;
- detecting parent and daughter ions by selected reaction monitoring (SRM);
- quantifying the concentration of said one or more free amino acids in accordance to the detected parent and/or daughter ions;
- and
- comparing levels of said one or more free amino acids in samples obtained before treatment to samples obtained after treatment.
A method of monitoring a subject's compliance with and response to dietary restriction due to a metabolic disorder, characterized by normalized levels of one or more free amino acids in samples obtained some time after onset of dietary restriction, the method comprising
- deproteinizing said samples to obtain free amino acids;
- separating the one or more free amino acids by liquid chromatography using strong ion-pairing conditions followed by weak ion-pairing conditions;
- ionizing said samples to generate ions;
- fragmenting ions to produce parent and daughter ions;
- detecting parent and daughter ions by selected reaction monitoring (SRM);
- quantifying the concentration of said one or more free amino acids in accordance to the detected parent and/or daughter ions;
- and
- comparing levels of said one or more free amino acids in samples obtained after onset of dietary restriction to samples obtained before onset of dietary restriction.
Dependencies are inferred from the claim text.