RRC ID 4059
著者 Kim JK, Harada K, Bamba T, Fukusaki E, Kobayashi A.
タイトル Stable isotope dilution-based accurate comparative quantification of nitrogen-containing metabolites in Arabidopsis thaliana T87 cells using in vivo (15)N-isotope enrichment.
ジャーナル Biosci Biotechnol Biochem
Abstract Stable isotope dilution-based comparative quantification of nitrogen-containing metabolites for highly sensitive and selective metabolomics was developed using liquid chromatography/mass spectrometry (LC/MS) and (15)N-isotope enrichment. We produced metabolically stable isotope-labeled Arabidopsis T87 cells by culturing with (15)N-labeled medium. We found that the growth of cells maintained in (15)N-labeled medium is very similar to the growth in normal medium, as evidenced by cell morphology, doubling time, and measurement of chlorophyll and carotenoid contents. Complete incorporation of (15)N in folate, S-adenosylmethionine (SAM), and S-adenosylhomocysteine (SAH) in T87 cells was accomplished after culturing for 21 d. Accurate comparative quantification of folate, SAM, and SAH was established by means of LC/MS using the isotopomers of the target metabolites as internal standards. The within- and between-run assay coefficients of variation for the folate, SAM, and SAH levels were all less than 8.5%. Stable isotope labeling by nitrogen source in Arabidopsis T87 cell culture provided simple, inexpensive, and accurate amino acid profiling. This interesting new protocol is valuable for the study of dynamic changes in N-compound pools in cultured cells.
巻・号 69(7)
ページ 1331-40
公開日 2005-7-1
DOI 10.1271/bbb.69.1331
PII JST.JSTAGE/bbb/69.1331
PMID 16041139
MeSH Arabidopsis / chemistry* Arabidopsis / cytology Chromatography, Liquid / methods Folic Acid / analysis Indicator Dilution Techniques / standards Mass Spectrometry / methods Nitrogen / analysis* Nitrogen / metabolism Nitrogen Isotopes Reproducibility of Results S-Adenosylhomocysteine / analysis S-Adenosylmethionine / analysis
IF 1.516
引用数 39
WOS 分野 CHEMISTRY, APPLIED FOOD SCIENCE & TECHNOLOGY BIOTECHNOLOGY & APPLIED MICROBIOLOGY BIOCHEMISTRY & MOLECULAR BIOLOGY
リソース情報
シロイヌナズナ / 植物培養細胞・遺伝子 rpc00008