Triglyceride assay by amperometric microbial biosensor: Sample hydrolysis and kinetic approach

A triglyceride assay based on triglyceride hydrolysis and glycerol detectio n was developed, Non-specific lipase isolated from Candida rugosa and intac t Gluconobacter oxydans cells, containing membrane-bound glycerol dehydroge nase, were used to develop a biosensor. Two approaches were investigated: a nalysis of pre-hydrolysed samples and a kinetic approach. The sensor prepar ed from G. oxydans cells exhibited sensitive and fast response to glycerol: detection limit 20 mu M (S/N=3), linear range up to 2 mM and response time 84 s (90% of steady-state). The triglyceride assay of pre-hydrolysed sampl es was based on a 20 min hydrolysis and determination of released glycerol by the biosensor. A calibration curve linear up to 12 mM was obtained for t riolein samples. The kinetic approach was based on simultaneous glyceride h ydrolysis and glycerol detection. Analysis time of 10 min, linear range up to 30 mM, and estimated detection limit of 50 mu M were achieved using the kinetic approach. The kinetic triglyceride assay is not influenced by free glycerol present in a sample. Storage stability, expressed as a half life ( 50 % of the initial response), was 7 days when trehalose was used as a stab iliser.