A number of assay protocols are employed for
lipase assay due to the wide substrate specificity oflipases. Determination of lipase activity at the lipidwater
interface is also indicative of free lipase. As
with all reactions catalyzed by enzymes, activity
measurements can be carried out using various
physico-chemical methods by monitoring the disappearance
of the substrate or by the release of the
product. Numerous methods are available for
measuring the hydrolytic activity as well as the
detection of lipase. The methods may be classified
as9: (i) Titrimetry, (ii) Spectroscopy (Photometry,
Fluorimetry and Infrared), (iii) Chromatography, (iv)
Radio activity, (v) Interfacial tensiometry, (vi) Turbidimetry,
(vii) Conductimetry, (viii) Immunochemistry,
and (ix) Microscopy. The general triacylglycerol
hydrolysis reaction catalyzed by lipases can be
written as
This reaction shows that the activity of lipases can
thus be assayed by monitoring the release of either
free fatty acids or glycerol from triacylglycerols or
fatty acid ester. The extensive reviews by Beisson
et al12 and Gupta et al9 describe in detail the various
methods available for lipase assay. Today the most
widely used lipase assay protocol is the titrimeteryassay using olive oil as a substrate because of its
simplicity, accuracy and reproducibility. A few
spectrophotometric assays are based on methods
which render colour to fatty acids released after
hydrolysis of triacylglycerols. A unit lipase activity13
relates to the release of 1 μmole of free fatty acid
from emulsified olive oil or triolein or tributyrin per
min at specified temperature and pH values. Specific
activity of lipases is expressed as units of lipolytic
activity per microgram of extra cellular protein.
A number of assay protocols are employed forlipase assay due to the wide substrate specificity oflipases. Determination of lipase activity at the lipidwaterinterface is also indicative of free lipase. Aswith all reactions catalyzed by enzymes, activitymeasurements can be carried out using variousphysico-chemical methods by monitoring the disappearanceof the substrate or by the release of theproduct. Numerous methods are available formeasuring the hydrolytic activity as well as thedetection of lipase. The methods may be classifiedas9: (i) Titrimetry, (ii) Spectroscopy (Photometry,Fluorimetry and Infrared), (iii) Chromatography, (iv)Radio activity, (v) Interfacial tensiometry, (vi) Turbidimetry,(vii) Conductimetry, (viii) Immunochemistry,and (ix) Microscopy. The general triacylglycerolhydrolysis reaction catalyzed by lipases can bewritten asThis reaction shows that the activity of lipases canthus be assayed by monitoring the release of eitherfree fatty acids or glycerol from triacylglycerols orfatty acid ester. The extensive reviews by Beissonet al12 and Gupta et al9 describe in detail the variousmethods available for lipase assay. Today the mostwidely used lipase assay protocol is the titrimeteryassay using olive oil as a substrate because of itssimplicity, accuracy and reproducibility. A fewspectrophotometric assays are based on methodswhich render colour to fatty acids released afterhydrolysis of triacylglycerols. A unit lipase activity13relates to the release of 1 μmole of free fatty acidfrom emulsified olive oil or triolein or tributyrin permin at specified temperature and pH values. Specificactivity of lipases is expressed as units of lipolyticactivity per microgram of extra cellular protein.
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