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2 protocols using 4 nitro 7 piperazino 2 1 3 benzoxadiazole nbd pz

1

Comprehensive Metabolic Analysis Protocol

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Lead nitrate (Pb(NO3)2), sodium lauryl sulfate, creatinine anhydrous, propionic acid, lithium d-, and l-lactate, acetylacetone, 37% formaldehyde, cimetidine, indoleacetic acid, methylumbelliferone, and methylumbelliferyl N-acetyl-β-d-glucosaminide were purchased from Sigma-Aldrich Fine Chemicals Inc. (MO, USA). Formic acid, HPLC grade acetonitrile (ACN), ethanol and methanol (MeOH) were purchased from Merck (Darmstadt, Germany). Hippuric acid, 2,2′-dipyridyl disulfate (DPDS), triphenylphosphine (TPP) and 4-nitro-7-piperazino-2,1,3-benzoxadiazole (NBD-PZ) were purchased from Tokyo Chemical Industry Co., Ltd. (Tokyo, Japan). 5-Aminolevulinic acid hydrochloride and guanidinoacetic acid were purchased from Alfa Aesar (MA, USA). Sulbactam was purchased from Cayman Chemical (Michigan, USA). Acetic acid was purchased from Hayashi Pure Chemical Ind.,Ltd. (Osaka, Japan). Trifluoroacetic acid (TFA) was purchased from Riedel-de Haën (Seelze, Germany). 3-(Trimethylsilyl)propionic-2,2,3,3-d4 acid sodium salt (TSP) was purchased from Cambridge Isotope Laboratories (MA, USA). 1,1-Dimethylbiguanide hydrochloride (metformin) was purchased from Santa Cruz Biotechnology, Inc. (Texas, USA). Protein Assay Dye Reagent was purchased from Bio-Rad Laboratories (California, USA).
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2

Enantioselective Derivatization and Quantification

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The enantiomers (sodium D-LA and L-LA) and a racemic mixture of sodium LA was purchased from Sigma Aldrich (St. Louis, MO, USA). Methanol (MeOH) and ethanol (EtOH) of HPLC grade, propionitrile (PrCN) and trifluoroacetic acid (TFA) were purchased from FUJIFILM Wako Pure Chemical Industries (Osaka, Japan). Acetonitrile (MeCN) of HPLC grade was obtained from Nacalai Tesque (Kyoto, Japan). 4-Nitro-7-piperazino-2,1,3-benzoxadiazole (NBD-PZ) and 4-(4,6-dimethoxy-1,3,5-triazin-2-yl)-4-methylmorpholinium chloride (DMT-MM) were products of Tokyo Chemical Industry (Tokyo, Japan). Water was purified by a Milli-Q Integral 3 system (Merck, Darmstadt, Germany). All other reagents were of the highest reagent grade and were used without further purification.
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