title: Development of quantitative enzymatic method and its validation for the assay of glucose in human serum.
abstract: OBJECTIVE
  To develop a simple, rapid, sensitive and affordable assay method for the determination of glucose in blood samples using a novel approach.
  
  
  METHODS
  A spectrophotometric method for glucose quantification in human serum samples based on self-coupling of activated 2,5-dimethoxyaniline (DMA) in the presence of peroxidase (POD)/glucose oxidase (GOD) and H(2)O(2) is described. H(2)O(2) generated in situ by catalytic reaction between GOD and glucose, activates DMA in the presence of POD to form a green-colored product, which has a strong absorption at λ(max)=740 nm at room temperature (30°C) in a 100 mmol/L acetate/acetic acid buffer of pH 4.2.
  
  
  RESULTS
  The linearity ranges for the quantification of glucose by rate and one-time detection method are 0.017-0.740 and 0.017-0.478 mmol/L, respectively. Within-day and day-to-day precision were 0.98-1.4% (n=10) and 1.33-2.89% (n=15), respectively. Glucose recoveries ranged from 96.6 to 102%, indicating minimal interference by commonly present interferants in serum samples. Accuracy results were between 90 and 102%. The detection and quantification limits of glucose were 2.376 and 7.923 μmol/L, respectively. The proposed method has good correlation coefficient of 0.999 with the enzymatic kit method.
  
  
  CONCLUSIONS
  This is a rapid and convenient method to determine serum glucose using simple spectrophotometer with excellent recovery and minimal interference by interferants in serum samples with low detection limit. Therefore, this method can be considered for adoption by the clinical diagnostic laboratories.
authors.affiliation: Department of Studies in Chemistry, University of Mysore, Manasagangotri, Mysore 570 006, Karnataka, India. profpn58@yahoo.com
authors.forename: Padmarajaiah
authors.initials: P
authors.lastname: Nagaraja
authors.lastname: Honnur Krishna
authors.forename: Anantharaman
authors.initials: A
authors.lastname: Shivakumar
authors.forename: Ashwinee K
authors.initials: AK
authors.lastname: Shrestha
mesh_terms: D000327:Adsorption; D000814:Aniline Compounds / Q000032:analysis; D001681:Biological Assay / Q000379:methods*; D001786:Blood Glucose / Q000032:analysis*; D002138:Calibration; D019411:Clinical Laboratory Techniques; D004305:Dose-Response Relationship, Drug; D005947:Glucose / Q000032:analysis*; D005949:Glucose Oxidase / Q000737:chemistry; D006801:Humans; D006861:Hydrogen Peroxide / Q000737:chemistry; D007700:Kinetics; D009195:Peroxidase / Q000737:chemistry; D011108:Polymers / Q000032:analysis; D015203:Reproducibility of Results; D044967:Serum / Q000378:metabolism*; D013696:Temperature
chemical_list: D000814:Aniline Compounds; D001786:Blood Glucose; D011108:Polymers; C542366:poly(2,5-dimethoxyaniline); D006861:Hydrogen Peroxide; D005949:Glucose Oxidase; D009195:Peroxidase; D005947:Glucose