---
title: Esculin hydrolysis test
description: "## About this test ### What is the purpose of the test?   The purpose is to see if the microbe can hydrolyze the compound esculin as a carbon source. ### How is esculin hydrolysis determined?  When microbes metabolize esculin…"
url: "https://vumicro.com/docs/esculin-hydrolysis-test/"
updated: "2022-11-01"
category: Reference Library / Biochemical tests
---

# Esculin hydrolysis test

## About this test

### What is the purpose of the test?  

The purpose is to see if the microbe can hydrolyze the compound esculin as a carbon source.

### How is esculin hydrolysis determined? 

When microbes metabolize esculin in a nutrient medium, the dark brown compound esculetin is produced and the medium is obviously discolored.

### What medium is used?  

Usually, the medium used is [bile esculin agar slant](https://vumicro.com/docs/bile-esculin-agar-slant/). This is a nutrient agar-based medium containing 0.1% esculin and 10% bile salts, and allowed to solidify at a slant. The bile salts inhibit some bacteria, and so the ability to grow in the presence of bile salts represents a second test use for the medium.

### How is the test performed?  

An inoculum from a pure culture is transferred aseptically to a sterile tube of [bile esculin agar](https://vumicro.com/docs/bile-esculin-agar-slant/) and streaked along the slant.   There is no need to stab the medium. The inoculated tube is incubated at 35-37 C for 24 hours and the results are determined. Abundant growth on the slant indicates a positive test for growth in the presence of bile. If growth is present, esculin hydrolysis can be observed if the medium has taken on an intense, chocolate brown coloration.

### What reagents are added?  

None.

## Performing this test in the VUMIE Online lab

### Inoculation of Medium

1. Select the [bile esculin agar slant](https://vumicro.com/docs/bile-esculin-agar-slant/) medium.  

2. Using an inoculating wire, complete the process of a [tube-to-tube aseptic transfer](https://vumicro.com/docs/perform-aseptic-transfers-tube-to-tube/) to inoculate the medium by streaking the slant and stabbing the butt of the medium.

### Incubation of the Inoculated Medium

3. Place the inoculated tube into the 35-37 C incubator.  

4. Press the [New Day button](https://vumicro.com/docs/use-the-new-day-button/) to move forward 24 hours.

### Determination of Test Results

5. Incubate this test for only 24 hours.  

6. Retrieve the incubated culture from the incubator.

7. Observe test result. The change of all or a significant portion of the medium to a chocolate brown indicates breakdown of esculin to esculetin, a positive test. Negative results would be indicated by growth on the slant but no change from the normal amber color of the medium.  

8. [Record test result](https://vumicro.com/docs/record-test-results/).  

[Uninoculated Bile Esculin Agar Slant](https://vumicro.com/wp-content/uploads/2022/07/ambysnnnnn-2018_07_01-17_21_39-UTC.jpg)

[Positive for Growth on Bile and Positive for Esculin Hydrolysis](https://vumicro.com/wp-content/uploads/2022/07/blaysynnnn-2018_07_01-17_21_39-UTC.jpg)

[Positive for Growth on Bile but Negative for Esculin Hydrolysis](https://vumicro.com/wp-content/uploads/2022/07/yeyysynnnn-2018_07_01-17_21_39-UTC.jpg)

[Negative for Growth on Bile](https://vumicro.com/wp-content/uploads/2022/07/ambysnnnnn-2018_07_01-17_21_39-UTC.jpg)

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