Expanded metabolite coverage of Saccharomyces cerevisiae extract through improved chloroform/methanol extraction and tert-butyldimethylsilyl derivatization
Artikel i vetenskaplig tidskrift, 2015

We present an improved extraction and derivatization protocol for GC-MS analysis of amino/non-amino acids in Saccharomyces cerevisiae. Yeast cells were extracted with chloroform: aqueous-methanol (1:1, v/v) and the resulting non-polar and polar extracts combined and dried for derivatization. Polar and non-polar metabolites were derivatized using tert-butyldimethylsilyl (t-BDMS) dissolved in acetonitrile. Using microwave treatment of the samples, the derivatization process could be completed within 2 h (from >20 h of the conventional method), providing fully derivatized metabolites that contain multiple derivatizable organic functional groups. This results in a single derivative from one metabolite, leading to increased accuracy and precision for identification and quantification of the method. Analysis of combined fractions allowed the method to expand the coverage of detected metabolites from polar metabolites i.e. amino acids, organic acids and non-polar metabolites i.e. fatty alcohols and long-chain fatty acids which are normally non detectable. The recoveries of the extraction method was found at 88 ± 4%, RSD, N = 3 using anthranilic acid as an internal standard. The method promises to be a very useful tool in various aspects of biotechnological applications i.e. development of cell factories, metabolomics profiling, metabolite identification, 13C-labeled flux analysis or semi-quantitative analysis of metabolites in yeast samples.

Metabolomics

Extraction

Derivatization

Saccharomyces cerevisiae

Författare

Sakda Khoomrung

Chalmers, Biologi och bioteknik, Systembiologi

Jose Luis Martinez Ruiz

Chalmers, Biologi och bioteknik, Systembiologi

Stefan Tippmann

Chalmers, Biologi och bioteknik, Systembiologi

Suwanee Jansa-Ard

Chalmers, Biologi och bioteknik, Systembiologi

Marieke Buffing

Chalmers, Biologi och bioteknik, Systembiologi

Raffaele Nicastro

Chalmers, Biologi och bioteknik, Systembiologi

Jens B Nielsen

Chalmers, Biologi och bioteknik, Systembiologi

Analytical Chemistry Research

2214-1812 (eISSN)

Vol. 6 9-16

Styrkeområden

Livsvetenskaper och teknik

Ämneskategorier

Bioinformatik och systembiologi

DOI

10.1016/j.ancr.2015.10.001