. 2018 Jan 20;28(3):167–179. doi: 10.1089/ars.2017.7014

Table 1.

Determination of NAD(P)(H) Interconversion Based on Spiking Extraction Solvent with Unlabeled Standards and Using it to Extract ¹³C-Labeled HCT116 Cells

Experiment	Species	Hot aqueous buffer	Enzyme assay buffer	Enzyme assay buffer with detergent	75% acetonitrile with KH₂PO₄	80% methanol	40:40:20 with 0.1 M FA	40:40:20 with 0.02 M FA	Standard alone
Spiking NADPH	NADPH	0.06 (±0.05)	0.44 (±0.05)	0.56 (±0.07)	0	0.65 (±0.11)	0.61 (±0.10)	0.69 (±0.05)	0.81 (±0.05)
	NADP⁺	0.20 (±0.15)	0.12 (±0.06)	0.04 (±0.03)	0.62 (±0.20)	0.06 (±0.04)	0.08 (±0.07)	0.03 (±0.02)	0.02 (±0.01)
Spiking NADP⁺	NADPH	0	0.14 (±0.05)	0.12 (±0.06)	0	0.02 (±0.01)	0.03 (±0.01)	0.01 (±0.01)	0
	NADP⁺	0.49 (±0.23)	0.53 (±0.09)	0.61 (±0.07)	0.88 (±0.15)	0.96 (±0.06)	0.88 (±0.07)	1.04 (±0.04)	1.23 (±0.05)
Spiking NADH	NADH	0.02 (±0.01)	0.48 (±0.16)	1.08 (±0.06)	0.12 (±0.11)	0.92 (±0.14)	0.41 (±0.09)	1.16 (±0.08)	1.34 (±0.07)
	NAD⁺	0.85 (±0.12)	0.56 (±0.13)	0.19 (±0.08)	0.84 (±0.18)	0.24 (±0.12)	0.54 (±0.13)	0.12 (±0.06)	0.03 (±0.02)
Spiking NAD⁺	NADH	0	0.54 (±0.08)	0.12 (±0.05)	0.21 (±0.13)	0.05 (±0.04)	0.13 (±0.06)	0.01 (±0.01)	0
	NAD⁺	0.73 (±0.14)	0.45 (±0.13)	1.04 (±0.12)	1.02 (±0.25)	1.06 (±0.05)	0.92 (±0.08)	1.20 (±0.08)	1.39 (±0.05)

Data are concentrations (μM) of the detected unlabeled species and are graphed in Figures 4 and 7.

There are small amounts of NADP⁺ contaminant detected in NADPH standard and NAD⁺ contaminant in NADH standard. The results are the average of three independent experiments and the error is the SD.

FA, formic acid; KH₂PO₄, monopotassium phosphate; NAD⁺, nicotinamide adenine dinucleotide; NADH, nicotinamide adenine dinucleotide reduced; NADP⁺, nicotinamide adenine dinucleotide phosphate; NADPH, nicotinamide adenine dinucleotide phosphate reduced; SD, standard deviation.