Reduced nicotinamide adenine dinucleotide phosphate

Reduced glutathione (GSH), oxidized glutathione (GSSG), glutathione reductase, γ-glutamylp-nitroanilide, glycylglycine, bovine serum albumin (BSA), 1,2-dithio-bis nitro benzoic acid (DTNB), 1-chloro-2,4-dinitrobenzene (CDNB), reduced nicotinamide adenine dinucleotide phosphate (NADPH), flavine adenine dinucleotide (FAD), glucose-6-phosphate, Tween-20, 2,6-dichlorophenolindophenol, thiobarbituric acid (TBA), picric acid, sodium tungstate, sodium hydroxide, trichloroacetic acid (TCA) and perchloric acid (PCA) were purchased from Sigma Chemicals Co. St. Louis, USA.

Emamectin Benzoate (EB) (~93.4%B1a, CAS: 155569-91-8, 98%), 7-benzyloxy-4-trifluoromethyl coumarin (BFC) and reduced nicotinamide adenine dinucleotide phosphate (NADPH) were purchased from Sigma-Aldrich company (St. Louis, Missouri, USA). Abamectin (B1a, CAS: 65195-55-3, 97%) was purchased from Toronto Research Chemicals (North York, Canada). Solvents for LC-MS (ammonium acetate, formic acid and acetonitrile) were obtained from Fisher Scientific (Pittsburgh, Pennsylvania, USA).
A series of gradient concentrations of insecticides diluted from stock solution with 0.1% (w/v) Triton X-100 water were prepared. In each well of the 24-well plate, a liquid artificial diet (~120 μL) was supplied. One hundred microliters of the insecticide solution was added to the dietary surface of every single well after the diet cooled and solidified. A third-instar larva was placed in each well. For each concentration, 24 larvae were treated. Triton X-100 (0.1%, w/v) was employed to process the control group. Two days later, larvae were checked and considered dead if they did not move after gentle prodding. The data were analyzed using DPS 7.05 software (Zhejiang University, Hangzhou, China). When the 95% fiduciary limits of the LC₅₀ values did not overlap, the LC₅₀ values were considered to vary significantly.

Racemic, (S)-(+)- and (R)-(−)- enantiomers of ¹³C(6)-labelled PQ (MW 259.1317 g/mol); PQ alcohol, 2-, 3-, and 4-hydroxyprimaquine and CPQ were synthesized as previously reported [9 (link), 22 (link)–24 (link)]. The identity of the compounds synthesized was confirmed by spectral infra-red (IR), nuclear magnetic resonance (NMR), high-resolution mass spectrometry and physical data in comparison with published values.
Co-factors for enzyme activity, including glucose-6-phosphate and its dehydrogenase, reduced nicotinamide adenine dinucleotide phosphate and magnesium chloride were purchased from Sigma-Aldrich (St Louis, MO, USA). Analytical grade solvents, including acetonitrile and methanol were purchased from Fisher Scientific (Fair Lawn, NJ, USA). Milli-Q system (Millipore, Bedford, MA, USA) was used to purify water for analytical utility.

Reduced glutathione (GSH), oxidized glutathione (GSSG), glutathione reductase, gamma-glutamyl p-nitroanilide, glycylglycine, bovine serum albumin (BSA), 1,2-dithio-bis nitro benzoic acid (DTNB), 1-chloro-2,4-dinitrobenzene (CDNB), reduced nicotinamide adenine dinucleotide phosphate (NADPH), CCl₄, flavine adenine dinucleotide (FAD), glucose-6-phosphate, Tween-20, 2,6-dichlorophenolindophenol, thiobarbituric acid (TBA), picric acid, sodium tungstate, sodium hydroxide, trichloroacetic acid (TCA) and perchloric acid (PCA) were purchased from Sigma Chemicals Co. USA.

The following reagents were used to synthesize, functionalize and conjugate gold nanostars: gold(III) chloride hydrate (HAuCl₄·xH₂O, 99.9%), silver nitrate (AgNO₃, 99.9%), HEPES (C₈H₁₈N₂O₄S, 99.5%), sodium chloride (NaCl, 99.0%), polyvinylpyrrolidone (PVP 10 000 (C₆H₉NO)_n, 99.0%), galactose (C₆H₁₂O₆, 99.0%), glucose (C₆H₁₂O₆, 99.0%), 3,3′-dithiobis(sulfosuccinimidyl propionate) (DTSSP, C₁₄H₁₄N₂O₁₄S₄Na₂), sodium hydroxide (NaOH, 98.0%), hydrogen peroxide (H₂O₂, 30% w/w), Tris buffer (NH₂C(CH₂OH)₃, 99.7%), galactose dehydrogenase (GalDH, approx. 80 U mg⁻¹ protein, 95.0%), aldose reductase (AR, 1 mg ml⁻¹, 95.0%), galactose oxidase (GalOx, 3000 U g⁻¹, 98.0%), reduced nicotinamide adenine dinucleotide phosphate (NADPH, C₂₁H₂₆N₇Na₄O₁₇P₃·xH₂O, 97.0%), reduced nicotinamide adenine dinucleotide (NAD, C₂₁H₂₇N₇Na₂O₁₄P₂·xH₂O, 97%) and nicotinamide adenine dinucleotide hydrate (NADH, C₂₁H₂₇N₇O₁₄P₂·xH₂O, 97.0%) were purchased from Sigma-Aldrich (Johannesburg, South Africa) and were used without any purification. Supplemented fetal bovine serum (Ham's F-12K) was purchased from Thermo Fisher Scientific (Johannesburg, South Africa). Blank urine was purchased from Industrial Analytical (Johannesburg, South Africa). Lastly, ultrapure water (ddH₂O, 18.2 MΩ cm⁻¹) was used for preparing all solutions from the high-purity chemicals and reagents.

Gallic acid, vanillin, acetylsalicylic acid, quercetin, rutin, 3,5-Di-tert-4-butylhydroxytoluene (BHT), ascorbic acid, sodium diclofenac, 1,1-Diphenyl-2-picrylhydrazyl (DPPH), guanidine hydrochloride,1-chloro-2,4-dinitrobenzene (CDNB), 5,50-dithiobis-2-nitrobenzoic acid (DTNB), reduced glutathione (GSH), oxidized GSH (GSSG), pyrogallol, 2,6-dichlorophenol-indophenol (DCPIP), potassium ferricyanide, triton X-100, ethylenediaminetetraacetic acid (EDTA), sodium pyruvate, thiobarbituric acid (TBA), reduced nicotinamide adenine dinucleotide (NADH) and reduced nicotinamide adenine dinucleotide phosphate (NADPH) were obtained from Sigma Chemical Co. (St. Louis, MO, USA). The rest of the chemicals used were procured from local firms (India) and were of highest purity grade.