title: Assay of serum arylesterase activity by fitting to the reaction curve with an integrated rate equation.
abstract: BACKGROUND
  Conventional enzyme activities make use of the initial reaction rate at high substrate concentrations. Because this is not always practical, alternative enzyme assays have been sought.
  
  
  METHODS
  Reaction curve fitting with an integrated rate equation was investigated to assay serum arylesterase (ArE) activity using phenyl acetate (PA) and p-nitrophenol acetate (PNPA) as substrates. At a much lower initial concentration of substrate (S(0)), the simplified integrated rate equation for the ArE reaction was ln(S(0)/S(i))=(V(m)/K(m)+K(d))t(i). Treating S(0) as a parameter, the enzyme activity as V(m)/K(m) was estimated through nonlinear least square fitting to reaction curve, and the multiplication of V(m)/K(m) by K(m) produced V(m). Spontaneous hydrolysis of the substrate with a rate constant, K(d), served as the background for the estimation of V(m)/K(m).
  
  
  RESULTS
  Substrate concentration at 8% of K(m) was well suited for the estimation of V(m)/K(m). With either substrate, the V(m)/K(m) showed a close relation to the percentage of substrate consumed, and was not affected by common systematic errors. With either substrate, the between-run precision for V(m)/K(m) was 6% (n>7), V(m)/K(m) was proportional to the amount of ArE and closely correlated with its initial rate. The upper limit of linearity by this integrated method was much higher than the initial rate method, while the detection limit was comparable. By using either V(m)/K(m) or the initial rate, there was negligible interference with ArE activity assay from triglycerides, bilirubin, and hemoglobin.
  
  
  CONCLUSIONS
  These results indicate the feasibility of the integrated method for routine assay of serum enzyme activity.
authors.affiliation: Biochemistry Department, Chongqing University of Medical Science, Chongqing 400016, PR China. liaofei@mail.cqums.edu.cn
authors.forename: F
authors.initials: F
authors.lastname: Liao
authors.forename: W L
authors.initials: WL
authors.lastname: Liu
authors.forename: Q X
authors.initials: QX
authors.lastname: Zhou
authors.forename: Z C
authors.initials: ZC
authors.lastname: Zeng
authors.forename: Y P
authors.initials: YP
authors.lastname: Zuo
mesh_terms: D000465:Algorithms; D001663:Bilirubin / Q000097:blood; D015415:Biomarkers; D002265:Carboxylic Ester Hydrolases / Q000097:blood*; D003627:Data Interpretation, Statistical; D006454:Hemoglobins / Q000032:analysis; D006801:Humans; D006868:Hydrolysis; D007700:Kinetics; D010648:Phenylacetates; D014280:Triglycerides / Q000097:blood
chemical_list: D015415:Biomarkers; D006454:Hemoglobins; D010648:Phenylacetates; D014280:Triglycerides; D002265:Carboxylic Ester Hydrolases; C019734:arylesterase; C025136:phenylacetic acid; C118008:4-nitrophenylacetic acid; D001663:Bilirubin