Gold (III) chloride trihydrate (product #: 520918) and sodium citrate dihydrate (product #: 567446) were purchased from Millipore Sigma. The NMR reference compound deuterated 3-1-propanesulfonic acid-d6 sodium salt (DSS-d6, DLM-8206), Tryptophan (15N labeled, NLM-800-PK), and 99.9% D2O (DLM-4) were purchased from Cambridge Isotope Labs (Tewksbury, MA). All chemicals were used as purchased without additional purification. Proteinase K (PK) was obtained from Amresco (#0706, biotechnology grade). Bovine serum albumin (BSA) was purchased from Calbiochem (# 12657, electrophoresis 100%). Human carbonic anhydrase II (HCA) was provided by Dr. Joseph P. Emerson at Mississippi State University. Before use, HCA was converted to apo-HCA by dialysis with ACES buffer containing dipicolinic acid at pH 7 described previously61 (link). p-Nitrophenyl acetate (pNPA) was purchased from Sigma-Aldrich (#N8130). All compounds purchased commercially were used without additional purification.
P nitrophenyl acetate p npa
Manufactured by Merck Group
Sourced in United States
P-nitrophenyl acetate (p-NPA) is a chemical compound commonly used as a substrate in various enzymatic assays. It is a colorless to pale yellow crystalline solid. p-NPA is often utilized to measure the activity of enzymes that catalyze the hydrolysis of esters, such as acetylcholinesterase and other esterases.
Automatically generated - may contain errors
Lab products found in correlation
Sodium citrate dihydrate, by Merck Group (1 mentions)
Gold 3 chloride trihydrate, by Merck Group (1 mentions)
Benzoic acid, by Merck Group (1 mentions)
Sucrose, by Merck Group (1 mentions)
Resazurin, by Merck Group (1 mentions)
Resorufin, by Merck Group (1 mentions)
Iodonitrotetrazolium chloride, by Merck Group (1 mentions)
Formic acid, by Merck Group (1 mentions)
Diaphorase from clostridium kluyveri, by Merck Group (1 mentions)
8 anilinonaphthalene 1 sulfonic acid, by Merck Group (1 mentions)
Bovine serum albumin (bsa), by Merck Group (1 mentions)
N acetyl l cysteine, by Merck Group (1 mentions)
Carbonic anhydrase, by Merck Group (1 mentions)
Methylglyoxal, by Merck Group (1 mentions)
Glyoxal, by Merck Group (1 mentions)
Transthyretin, by Merck Group (1 mentions)
2 thiobarbituric acid, by Merck Group (1 mentions)
Dinitrophenylhydrazine, by Merck Group (1 mentions)
P nitrophenol pnp, by Merck Group (1 mentions)
Triton x 100, by Merck Group (1 mentions)
8 protocols using p nitrophenyl acetate p npa
1
Preparation of Apo-HCA for Enzymatic Assays
2
PET Film Characterization and pNPA Hydrolysis
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PET film (ES301445, amorphous, transparent, 0.25 mm thickness) was purchased from Goodfellow GmbH (London, United Kingdom). BHET (> 85%) was purchased from Tokyo Chemical Industry (Shanghai, China). p-Nitrophenyl acetate (pNPA) was purchased from Sigma-Aldrich (St. Louis, MO, United States). Other chemicals (Supplementary Table 2 ) used in this study were of analytical grade and purchased from commercial sources.
3
Cutinase Cut190 S184P/R186S Catalysis
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Genes coding for the cutinase Cut190 S184P/R186S (S226P/R228S including the signal peptide, GenBank BAO42836.1), kREDy‐to‐goTM plates and KREDs 54, 61, 136, 181, 213 and 296 were kindly provided by Prozomix Ltd. Escherichia coli BL21(DE3) competent cells were purchased from Stratagene.
Benzoic acid, bis(2‐hydroxylethyl) terephthalate (BHET), diaphorase from Clostridium kluyveri, formic acid, iodonitrotetrazolium chloride (INT), pnitrophenyl acetate (pNPA), resazurin and resorufin were purchased from Sigma (San Louis, USA). β‐Nicotinamide adenine dinucleotide (NAD+), β‐nicotinamide adenine dinucleotide phosphate (NADP+), β‐nicotinamide adenine dinucleotide reduced form (NADH) and β‐nicotinamide adenine dinucleotide phosphate reduced form (NADPH) were from Carbosynth Ltd. (Compton, UK). Benzonase nuclease was from Novagen (Madison, USA). Ethylene glycol (EG) was from May & Baker Ltd. (Dagenham, United Kingdom). Mono(2‐hydroxyethyl) terephthalate (MHET) was from Activate Scientific GmbH (Prien am Chiemsee, Germany). Sucrose was from Merck Millipore Ltd. PET films (250 μm thickness, transparent, amorphous) were purchased from GoodFellow Inc. (Wrexham, UK).
Benzoic acid, bis(2‐hydroxylethyl) terephthalate (BHET), diaphorase from Clostridium kluyveri, formic acid, iodonitrotetrazolium chloride (INT), pnitrophenyl acetate (pNPA), resazurin and resorufin were purchased from Sigma (San Louis, USA). β‐Nicotinamide adenine dinucleotide (NAD+), β‐nicotinamide adenine dinucleotide phosphate (NADP+), β‐nicotinamide adenine dinucleotide reduced form (NADH) and β‐nicotinamide adenine dinucleotide phosphate reduced form (NADPH) were from Carbosynth Ltd. (Compton, UK). Benzonase nuclease was from Novagen (Madison, USA). Ethylene glycol (EG) was from May & Baker Ltd. (Dagenham, United Kingdom). Mono(2‐hydroxyethyl) terephthalate (MHET) was from Activate Scientific GmbH (Prien am Chiemsee, Germany). Sucrose was from Merck Millipore Ltd. PET films (250 μm thickness, transparent, amorphous) were purchased from GoodFellow Inc. (Wrexham, UK).
4
Esterase Activity Determination Protocol
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All the chemicals used in the present investigation were procured from Hi-Media and Merck, Mumbai, India. The substrate p-nitrophenyl acetate (p-NPA) for esterase assay was procured from Sigma, Aldrich (U.S.A.) .
5
Carbonic Anhydrase Inhibition Assay
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Carbonic anhydrase (CA, from bovine erythrocytes), Bovine serum albumin (BSA, from bovine), Transthyretin (TTR, from human plasma), and other chemicals were purchased from Sigma-Aldrich chemical Co. S-allyl l -cysteine (SAC) was purchased from Cayman Chemicals. MethylGlyoxal (MGO), Glyoxal (GO), Fructose, 8-Anilinonaphthalene-1-sulfonic acid (ANS), Dinitrophenyl hydrazine (DNPH), p-Nitrophenyl acetate (p-NPA), 2-Thiobarbituric acid (TBA) and N-acetyl l -cysteine (NAC) were also obtained from Sigma Chemical Co.
6
Lipase Activity Assay using p-NPP
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For lipase assay, the substrate solution was prepared by adding 100 µL of the p-NPP solution (7.5 mg p-NPP from Sigma Aldrich, Italy, dissolved in 2 mL of propane-2-ol and acetonitrile 1:1) to 800 µL of 0.05 mol L -1 Tris-HCl buffer (pH 8.0) and 75 µL of water. The lipase assay was carried out at 25 °C in the presence of Triton X-100 (20 µl mL -1 ) [27] (link) by adding 25 µL of the enzyme extract to the substrate solution. After 20 min, the reaction was stopped by incubating the mixture on ice. The absorbance was spectrophotometrically measured at 405 nm, against an enzyme-free control.
One lipase unit (U) was defined as the amount of enzyme that liberates 1 µmol of p-nitrophenol per min, under the described assay conditions. All enzyme assays were carried out in double and the average values were calculated.
The eventual unspecific esterase activity was spectrophotometrically determined following the protocol described above but replacing the p-nitrophenyl palmitate with p-nitrophenyl acetate (p-NPA) (Sigma Aldrich, Italy) at 25 °C and pH 8.0.
One lipase unit (U) was defined as the amount of enzyme that liberates 1 µmol of p-nitrophenol per min, under the described assay conditions. All enzyme assays were carried out in double and the average values were calculated.
The eventual unspecific esterase activity was spectrophotometrically determined following the protocol described above but replacing the p-nitrophenyl palmitate with p-nitrophenyl acetate (p-NPA) (Sigma Aldrich, Italy) at 25 °C and pH 8.0.
7
PET Substrate Characterization Protocol
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PET fiber was kindly provided by Lealea Enterprise (Taipei, Taiwan) and high-crystallinity PET was obtained from drinking water bottle. Surface area of the PET fibers and PET bottle powder were analyzed using surface area and pore size analyzer (BEL Japan, Inc.). The crystallinity of PET substrates was analyzed using X-Ray Diffraction (XRD) (D2 phaser, Bruker, Germany). Acetonitrile of HPLC grade was obtained from Merck. p-Nitrophenol (pNP) and p-nitrophenyl acetate (pNPA) were obtained from Sigma-Aldrich (USA) and Alfa Aesar (Massachusetts, USA), respectively. All other reagents used were analytical grade available from Acros, Merck, and Sigma-Aldrich.
8
Thiamethoxam Toxicity Assay Protocol
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Thiamethoxam (98.5% pure) was purchased from Dr. Ehrenstorfer GmbH. Yeast extract, D-glucose, D-fructose, antipain, aprotinin, leupeptin, pepstatin A, soybean trypsin inhibitor, monosodium phosphate, sodium chloride (NaCl), Triton ® X-100, acetylthiocholine iodide (AcSCh.I), 5,5'dithio-bis(2,nitrobenzoic acid) (DTNB), reduced L-glutathione (GSH), ethylenediaminetetraacetic acid (EDTA), 1-chloro-2,4-dinitrobenzene (CDNB), Trizma ® base (Tris), hydrochloric acid (HCl), magnesium chloride (MgCl 2 ), pnitrophenyl phosphate (p-NPP), 1,5-bis(4allyldimethylammonium-phenyl)pentan-3-one-dibromide (BW284C51) and p-nitrophenyl acetate (p-NPA) were obtained from Sigma Aldrich (France). Royal jelly was purchased from Ickowicz Apiculture (Boll ene, France).
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