Darkening
Media containing sugars and peptones undergo the Maillard reaction at 121 °C, so a slightly deeper colour than the freshly prepared solution is normal. If it turns distinctly brown:
- The cycle was too long or too hot. Large vessels heat slowly and are often over-sterilised by ten minutes or more.
- High-sugar formulations such as Sabouraud and Potato Dextrose are more sensitive; the specification sheet may call for 115 °C or a shorter cycle.
- Sterilising sugar and peptone separately and combining afterwards is the standard approach for heat-sensitive formulations.
Turbidity or precipitate
- Phosphate precipitation. Formulations containing both phosphate and magnesium or calcium can throw a white precipitate after autoclaving. Dissolving the phosphate first, or sterilising the salts separately, helps.
- Tap water. Hardness and calcium or magnesium ions cause turbidity; use purified water.
- Protein precipitation at low pH. Check the preparation pH.
- Undissolved agar. See "Why is my agar medium soft or not setting?".
- Black precipitate. Usually linked to iron and sulphur-containing components; see "Why does fermentation broth turn black after sterilisation?".
When to discard
Obvious caramelisation, a burnt smell, a precipitate that does not disperse on swirling, or poorer growth of control strains all mean the batch should not be used for routine testing. Minor colour differences with normal performance testing are acceptable.