Evaluation of lophine derivatives as L-012 (luminol analog)-dependent chemiluminescence enhancers for measuring horseradish peroxidase and H2O2

• Published in: Luminescence (Chichester, England) Vol. 29; no. 2; pp. 118 - 121
• Main Authors: Ichibangase, T., Ohba, Y., Kishikawa, N., Nakashima, K., Kuroda, N.
• Format: Journal Article
• Language: English
• Published: England Blackwell Publishing Ltd 01.03.2014; Wiley Subscription Services, Inc
• Subjects: chemiluminescence enhancer; Horseradish Peroxidase - analysis; Horseradish Peroxidase - metabolism; HRP; Hydrogen Peroxide - analysis; Imidazoles - chemistry; lophine derivative; Luminescence; Luminescent Agents - chemical synthesis; Luminescent Agents - chemistry; Luminol - analogs & derivatives; Luminol - chemical synthesis; Luminol - chemistry; luminol analog; Molecular Structure; luminol analog; chemiluminescence enhancer; HRP; lophine derivative
• ISSN: 1522-7235; 1522-7243
• DOI: 10.1002/bio.2513

ABSTRACT8‐Amino‐5‐chloro‐7‐phenylpyrido[3,4‐d]pyridazine‐1,4(2H,3H)dione (L‐012) was recently synthesized as a new chemiluminescence (CL) probe; the light intensity and the sensitivity of L‐012 are higher than those of other CL probes such as luminol. Previously, our group developed four lophine‐based CL enhancers of the horseradish peroxidase (HRP)‐catalyzed CL oxidation of luminol, namely 2‐(4‐hydroxyphenyl)‐4,5‐diphenylimidazole (HDI), 2‐(4‐hydroxyphenyl)‐4,5‐di(2‐pyridyl)imidazole (HPI), 4‐(4,5‐diphenyl‐1H‐imidazol‐2‐yl)phenylboronic acid (DPA), and 4‐[4,5‐di(2‐pyridyl)‐1H‐imidazol‐2‐yl]phenylboronic acid (DPPA), and showed that DPPA was suitable for the photographic detection of HRP. In this study, we replaced luminol with L‐012 and evaluated these as L‐012‐dependent CL enhancers. In addition, to detect HRP and/or H2O2 with higher sensitivity, each detection condition for the L‐012–HRP–H2O2 enhanced CL was optimized. All the derivatives enhanced the L‐012‐dependent CL as well as luminol CL; HPI generated the highest enhanced luminescence. Under optimized conditions for HRP detection, the detection limit of HRP was 0.08 fmol. By contrast, the detection limit of HRP with the enhanced L‐012‐dependent CL using 4‐iodophenol, which is a common enhancer of luminol CL, was 1.1 fmol. With regard to H2O2 detection, the detection limits for enhanced CL with HPI and 4‐iodophenol were 0.29 and 1.5 pmol, respectively. Therefore, it is demonstrated that HPI is the most superior L‐012‐dependent CL enhancer. Copyright © 2013 John Wiley & Sons, Ltd. Copyright © 2013 John Wiley & Sons, Ltd.