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オービトラップ質量分析計を用いたアミノ酸解析の可能性
島村, 繁;澄田, 智美;布浦, 拓郎
Permalink : http://hdl.handle.net/2115/81350
JaLCDOI : 10.14943/lowtemsci.79.37
KEYWORDS : アミノ酸;同位体;キャピラリー電気泳動;質量分析;Amino acid;Stable isotope;Capillary electrophoresis;Mass spectrometry
Abstract
キャピラリー電気泳動インターフェイス（ZipChipTM）を備えたオービトラップ搭載質量分析計（Orbitrap Fusion）をアミノ酸分析へ適用すると，ガスクロマトグラフ-質量分析計（GC-MS）での分析と同程度あるいはそれ以下の試料量でも測定が可能なだけでなく，非常に簡便かつ短時間で解析することができる．更に，菌体を超音波破砕し得られた溶液に含まれる遊離アミノ酸を，ほとんど未調整のままで測定する事も可能である．これによりGC-MS 法における技術的課題：試料調製における（1）加水分解過程でのアスパラギン，グルタミンからのアスパラギン酸，グルタミン酸への変換（2）試料調製におけるプロリンの低い回収率，（3）GC-MS 解析におけるアルギニン，システイン，ヒスチジン，トリプトファンの検出不可，などの改善が期待される．本報では一般的なバクテリア培養菌体を用いて，遊離アミノ酸と総アミノ酸，それぞれの基本的な試料調整方法や測定条件等を紹介する．
Combination of a capillary electrophoresis interface ZipChipTMand Orbitrap Fusion mass spectrometer (Fusion MS) is applicable to amino acid analysis even from tiny amounts of biological samples, in the similar sensitivity to the conventional method using a gas chromatograph-mass spectrometer (GC-MS). Moreover, Fusion MS analysis can achieve simple preparation of samples and rapid detection of amino acids, which allow us to access cellular free amino acids with almost no preparation. Fusion MS analysis indeed potentially resolve the technical issues associated with the conventional GC-MS analysis, such as (1) conversion of asparagine and glutamine to asparaginate and glutamate, respectively, in protein hydrolysis and (2) low recovery of proline in sample preparation and no detection of arginine, cysteine, histidine, and tryptophan in GC-MS analysis. Here, we introduce sample preparation procedure for the analysis of free and total amino acids in bacterial cells and analytical conditions for the amino acid measurement in the Fusion MS method.
Combination of a capillary electrophoresis interface ZipChipTMand Orbitrap Fusion mass spectrometer (Fusion MS) is applicable to amino acid analysis even from tiny amounts of biological samples, in the similar sensitivity to the conventional method using a gas chromatograph-mass spectrometer (GC-MS). Moreover, Fusion MS analysis can achieve simple preparation of samples and rapid detection of amino acids, which allow us to access cellular free amino acids with almost no preparation. Fusion MS analysis indeed potentially resolve the technical issues associated with the conventional GC-MS analysis, such as (1) conversion of asparagine and glutamine to asparaginate and glutamate, respectively, in protein hydrolysis and (2) low recovery of proline in sample preparation and no detection of arginine, cysteine, histidine, and tryptophan in GC-MS analysis. Here, we introduce sample preparation procedure for the analysis of free and total amino acids in bacterial cells and analytical conditions for the amino acid measurement in the Fusion MS method.
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