**2.4.1 Materials required antimicrobial activity**

Bacterial culture; Mueller Hinton agar medium; Nutrient broth; Plant extracts; Bunsen flame; Micro pipettor (10µl-200µl), adjustable; Spreader/wire loop; Agar borer and Incubator.

#### **2.4.2 Method antimicrobial activity**

Approximately 2-5 freshly grown bacterial colonies of the test organism were emulsified into Nutrient broth and incubated for 10-15 minutes at room temperature. With a spreader of wire loop, Mueller Hinton agar plate was evenly inoculated and plate allowed to stand for 5 minutes at room temperature (i.e. 25oC).

Using a sterile agar borer (Sterilized using a bunsen flame), wells were dug into the inoculated agar at reasonable distance apart (approximately 5 cm).

The plant extract(s) were transferred into the created agar wells till when full. Extract were not allowed to float on the agar surface. The plate lid was replaced and did not turn the petri-dish upside down. The setup was incubated at 37oC overnight. The presence for bacterial inhibition zones around each well looked for. Appearance of clear zones around a well was indicative of the anti bacterial activity of an extracts (Bizimenyera *et al*., 2005).
