How to Test Food for Enzymes

Enzyme testing reveals important quality markers for a food product, like the degree to which the product is processed or the lack of processing altogether. Testing for enzymes in your product is imperative because untreated enzymes often lead to issues with food quality. If your product is treated to intentionally denature enzymes, or is a processed food that uses enzymes as an aid, the enzyme level should be tested to determine over- or under-use.

Armed with these understandings and those to come in this article, you’ll be well-equipped to proceed with enzyme testing for your food products.

What You Should Know About Enzymes and Testing

Before conducting an enzyme test, there are a few things that are beneficial to understand.

Unlike many food tests, enzyme results require context to understand them. It can be difficult to draw any conclusions from a single laboratory result for a number of reasons.

Not all labs conduct the same enzyme tests or use identical methods. That means reporting units may not be equivalent or comparable. Ideally, you’ll want to turn to a consistent partner for enzyme testing. Additionally, enzyme test results may be best viewed as a comparison before and after processing.

Interestingly, enzymes do not have a single source. They are derived from a variety of materials, including plant-based, mammalian, fungal, and bacterial. Because the sources are different, the enzymes may also present different activities and deactivation needs.

Medallion Labs tests enzymes at specific pH and temperatures as outlined for each assay. These may not match your real-world food processing methods. This is why comparative test results often have more value than an individual test result.

Understanding Enzyme Tests and Results

Medallion Labs offers 10 unique enzyme testing options. Each one demonstrates the activity of the enzyme for that specific assay. Enzyme substrates can be specific or generalized and may require processing for the enzymes to be active on them.

Which enzyme test will best answer your question? Review our menu of enzyme testing options and the short description of each one. If you need additional information, use the link to visit each test's specific page. We will note here that only two of the methods have any affiliation: Alpha-Amylase = AOAC 2002.01 and Trypsin Inhibitor = AACC 22-40.

Alpha-Amylase Activity Test

The Alpha-Amylase Activity Test is the most requested enzyme assay from Medallion Labs. Extracted samples are reacted with an alpha-amylase-specific substrate. Colored product is released by amylase and measured using a spectrophotometer.

Alpha-Amylase Inhibitor Test

Porcine pancreatic alpha-amylase is pre-incubated with four different dilutions of the sample in a 50 mM maleate buffer, pH 6.0, at 40°C for 20 minutes prior to alpha-amylase activity determination and evaluated against a control pre-incubated with buffer alone. Alpha amylase activity was assayed by AOAC official method 2002.01.

One alpha-amylase inhibitor unit (AAIU) is defined as the amount that will reduce two units of alpha-amylase by 50% under standard assay conditions.

Beta-Amylase Test

Extracted samples are reacted with a beta-amylase-specific substrate. Colored product is released by beta-amylase and measured on a spectrophotometer.

Beta-Glucanase Activity Test

Extracted samples are reacted with a beta-glucanase-specific substrate. A colored product is released, and beta-glucanase activity is measured on a spectrophotometer.

Lipase Activity Test

Samples are homogenized in a cold phosphate-buffered saline solution (PBS), pH 8.5 and then centrifuged. A protein determination by the Bradford method is performed on the enzyme extract (supernatant) to normalize subsequent enzyme results to the efficiency of the protein extraction. The enzyme extract is incubated at 37°C with a saturating amount of triglyceride that contains sulfhydryl groups substituted for the normal carboxylic acid groups on the fatty acids. As the extracted lipase in the supernatant cleaves the modified fatty acids from the glycerol backbone, the sulfhydryl groups are made available for reaction with a color reagent (DTNB). The color reagent absorbs visible light at 412nm in direct proportion to the number of sulfhydryl groups exposed by the lipase. The appearance of absorbance with time of reaction is monitored kinetically. The initial velocity of the reaction is determined from the reaction rate.

Lipoxygenase Activity Test

Samples are homogenized, and enzymes are extracted into a cold, pH 6.5 buffer. The enzyme extract is mixed with linoleic acid or arachidonic acid. Lipoxygenase activity is measured spectrophotometrically by the rate of formation of the fatty acid hydroperoxide at 234nm.

Peroxidase Activity Test

Samples are homogenized, and enzymes are extracted into a cold, pH 6.5 buffer. A protein determination by the Bradford method is performed on the enzyme extract (supernatant) to normalize subsequent enzyme results to the efficiency of the protein extraction. The enzyme extract is mixed with hydrogen peroxide, guaiacol, and calcium. Peroxidase activity is measured spectrophotometrically by the rate of formation of tetraguaiacol at 470nm.

Polyphenol Oxidase Activity Test

Samples are homogenized and enzymes are extracted into a cold, pH 6.5 buffer. The enzyme extract is mixed with catechol and polyphenol oxidase activity is measured spectrophotometrically by the rate of formation of 1,2 benzoquinone at 410nm.

Protease Activity Test

Extracted samples are reacted with a protease-specific substrate. Colored product is released, and protease activity is measured on a spectrophotometer.

Trypsin Inhibitor Activity Test

Samples are extracted in a buffered solution. Five dilutions are made, and trypsin substrate is added. Trypsin is then added at timed intervals. After 10 minutes of incubation, the reaction is stopped, and a colored product is produced. Any trypsin inhibitor present reduces the colored product produced.

Our Experts Make Enzyme Testing Successful

When considering Medallion Labs for enzyme testing or other lab services, we suggest referring to our tech data sheets and online order form for the latest details. Our breadth of testing options helps you maximize your testing budget by providing the results you seek in a cost-efficient manner.

Medallion Labs is ready to use our testing options, expertise, and skills for your testing needs. Connect with us or call 1-800-245-5615 to discuss the criteria and testing needs for your food products.