In the reference method described, what is the substrate used?

• A. Triolein
• B. Olive oil
• C. Glycerol
• D. Fatty acid

Answer: (A)

In this reference method, the substrate is the compound the enzyme acts upon to produce measurable products, so it must be a well-defined triglyceride for consistent results. Triolein is used because it is a single, defined triglyceride (glycerol esterified with three oleic acid molecules), providing a stable and standardized substrate. Olive oil, while containing triglycerides, is a mixture with variable composition and isn’t ideal for standardization. Glycerol and fatty acids are the hydrolysis products, not the starting substrate. Therefore, triolein is the substrate chosen.