Application of cyanide hydrolase from Klebsiella sp. in a biosensor system for the detection of low-level cyanide.
A partially purified preparation of cyanide hydrolase (cyanidase) from a bacterium, Klebsiella sp., was applied as a biocatalyst in a biosensor system for low-level cyanide detection. In the biosensor system cyanide hydrolase converts cyanide into formate and ammonia. The formate produced in the cyanide degradation was detected with a formate biosensor, in which formate dehydrogenase (FDH; E.C. 1.2.1.2) was co-immobilized with salicylate hydroxylase (SHL; E.C. 1.14.13.1) on a Clark electrode. The principle of the formate sensor is that FDH converts formate into carbon dioxide using beta-nicotinamide adenine dinucleotide hydrate (NAD(+)). The corresponding NADH produced is then oxidized to NAD(+) by SHL using salicylate and oxygen. The oxygen consumption is monitored with the Clark electrode. The optimum buffer pH and temperature for the enzymatic hydrolysis of potassium cyanide were studied. The preliminary experiments including the pretreatment of cyanide with cyanide hydrolase and then detection by the formate sensor gave a detection limit at 7.3 micromol l(-1) cyanide. The linear range of the calibration curve was between 30 micromol l(-1) and 300 micromol l(-1) cyanide.
Mak
Karen K W
KK
Biosensors and Bioelectronics Laboratory, Department of Chemistry, The Hong Kong University of Science and Technology, Clear Water Bay, Kowloon, Hong Kong, China. karen@ust.hk
Law
Alex W C
AW
Tokuda
Shinsuke
S
Yanase
Hideshi
H
Renneberg
Reinhard
R
D000641:Ammonia / Q000378:metabolism; D015374:Biosensing Techniques / Q000379:methods*; D002245:Carbon Dioxide / Q000378:metabolism; D003486:Cyanides / Q000032:analysis*; D005560:Formate Dehydrogenases / Q000378:metabolism; D005561:Formates / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D006867:Hydrolases / Q000378:metabolism*; D007709:Klebsiella / Q000201:enzymology*; D006899:Mixed Function Oxygenases / Q000378:metabolism; D009243:NAD / Q000378:metabolism; D010100:Oxygen / Q000378:metabolism; D010101:Oxygen Consumption; D020156:Salicylic Acid / Q000378:metabolism; D012680:Sensitivity and Specificity; D013696:Temperature
D003486:Cyanides; D005561:Formates; D009243:NAD; C030544:formic acid; D002245:Carbon Dioxide; D000641:Ammonia; D006899:Mixed Function Oxygenases; C022619:salicylate 1-monooxygenase; D005560:Formate Dehydrogenases; D006867:Hydrolases; C070397:cyanidase; D020156:Salicylic Acid; D010100:Oxygen