PMID- 37651912
OWN - NLM
STAT- MEDLINE
DCOM- 20230915
LR  - 20230915
IS  - 1873-3573 (Electronic)
IS  - 0039-9140 (Linking)
VI  - 266
IP  - Pt 2
DP  - 2024 Jan 1
TI  - Dual strategy for (13)C-Metabolic flux analysis of central carbon and energy 
      metabolism in Mammalian cells based on LC-isoMRM-MS.
PG  - 125074
LID - S0039-9140(23)00825-1 [pii]
LID - 10.1016/j.talanta.2023.125074 [doi]
AB  - Central carbon and energy metabolism are the most concerned metabolic pathways in 
      (13)C-Metabolic flux analysis ((13)C-MFA). However, some alpha-keto acids, 
      ribonucleoside triphosphate (NTPs) and deoxyribonucleoside triphosphate (dNTPs) 
      involved in central carbon and energy metabolism pathways were unstable or 
      reactive, leading to inaccurate metabolic flux analysis. To achieve accurate 
      (13)C-MFA of central carbon and energy metabolism, we proposed a dual strategy 
      for the detection of 101 metabolites in glucose metabolism pathways. 
      N-Methylphenylethylamine (MPEA) was utilized for derivatization of 4 carboxyl 
      (alpha-keto acids) and 8 phosphate metabolites (NTPs and dNTPs). After 
      derivatization, the MPEA derivatives were investigated to be stable for 4 weeks 
      under 4  degrees C and detected with high intensity in  approximately 10(4) cells. On the other hand, 
      we analyzed an additional 89 metabolites in central carbon and energy metabolic 
      pathways were directly analyzed by liquid chromatography tandem mass spectrometry 
      (LC-MRM-MS). The limit of detection (LODs) of our method were as low as 
      0.05 ng/mL and the linear range was at least two orders of magnitude with 
      determination coefficient (R(2)) > 0.9701. The relative standard divisions (RSDs) 
      of intra- and inter-day of 95% metabolites were below 20%. In addition, the 
      isotope list of 82 detected metabolites in central carbon and energy metabolism 
      were generated according to isotopologues and isotopomers for each metabolite 
      resulting from (13)C incorporation. Accurate assessment of mass isotopomer 
      distributions (MIDs) of intracellular (13)C-labeled metabolites was achieved in 
      [U-(13)C]-glucose cultured HepG2 cells by our dual strategy. Finally, we 
      performed MID analysis of 101 metabolites in central carbon and energy 
      metabolism. Overall, this dual method is reproducible and robust for application 
      on (13)C-MFA and has a great potential for studying clinical isotope labeled 
      samples.
CI  - Copyright (c) 2023 Elsevier B.V. All rights reserved.
FAU - Zheng, Jie
AU  - Zheng J
AD  - Singapore Phenome Center, Nanyang Technological University, 639798, Singapore.
FAU - Yang, Junjie
AU  - Yang J
AD  - School of Civil and Environmental Engineering, Nanyang Technological University, 
      639798, Singapore; Nanyang Environment & Water Research Institute, Nanyang 
      Technological University, 637141, Singapore.
FAU - Liang, Xu
AU  - Liang X
AD  - Singapore Phenome Center, Nanyang Technological University, 639798, Singapore.
FAU - Fang, Mingliang
AU  - Fang M
AD  - School of Civil and Environmental Engineering, Nanyang Technological University, 
      639798, Singapore; Department of Environmental Science and Engineering, Fudan 
      University, Shanghai, 200433, China. Electronic address: mlfang@ntu.edu.sg.
FAU - Wang, Yulan
AU  - Wang Y
AD  - Singapore Phenome Center, Nanyang Technological University, 639798, Singapore; 
      Lee Kong Chian School of Medicine, Nanyang Technological University, 639798, 
      Singapore. Electronic address: liang.xu@ntu.edu.sg.
LA  - eng
PT  - Journal Article
DEP - 20230822
PL  - Netherlands
TA  - Talanta
JT  - Talanta
JID - 2984816R
RN  - NU43IAG5BC (triphosphoric acid)
RN  - 7440-44-0 (Carbon)
RN  - 0 (Keto Acids)
RN  - 0 (Polyphosphates)
SB  - IM
MH  - Animals
MH  - *Metabolic Flux Analysis
MH  - *Energy Metabolism
MH  - Carbon
MH  - Chromatography, Liquid
MH  - Keto Acids
MH  - Polyphosphates
MH  - Mammals
OTO - NOTNLM
OT  - (13)C-metabolic flux analysis
OT  - Carboxyl and phosphate metabolites
OT  - Central carbon and energy metabolism
OT  - Derivatization
OT  - Liquid chromatography-mass spectrometry
OT  - Mass isotopomer distributions
COIS- Declaration of competing interest The authors declare that they have no known 
      competing financial interests or personal relationships that could have appeared 
      to influence the work reported in this paper.
EDAT- 2023/09/01 00:41
MHDA- 2023/09/15 06:42
CRDT- 2023/08/31 18:08
PHST- 2023/06/01 00:00 [received]
PHST- 2023/08/06 00:00 [revised]
PHST- 2023/08/10 00:00 [accepted]
PHST- 2023/09/15 06:42 [medline]
PHST- 2023/09/01 00:41 [pubmed]
PHST- 2023/08/31 18:08 [entrez]
AID - S0039-9140(23)00825-1 [pii]
AID - 10.1016/j.talanta.2023.125074 [doi]
PST - ppublish
SO  - Talanta. 2024 Jan 1;266(Pt 2):125074. doi: 10.1016/j.talanta.2023.125074. Epub 
      2023 Aug 22.