Overview:
Antibiotic Medium No. 9 is the official standardized base-layer agar used in the cylinder-plate microbiological assay of polymyxin B and colistin (polymyxin E). It is designated as �Medium I� in USP <81>, European Pharmacopoeia 2.7.2, JP 17, and WHO guidelines. When overlaid with a seed layer of Antibiotic Medium No. 10 inoculated with Bordetella bronchiseptica, it produces exceptionally large, sharp, and highly reproducible inhibition zones specific to polymyxins. The medium is enriched and slightly more alkaline than general-purpose base agars to maximize polymyxin diffusion.
Applications:
?? Compendial base layer for polymyxin B sulfate and colistin sulfate potency assays
?? Cylinder-plate diffusion assay of polymyxin-containing pharmaceuticals
?? Regulatory QC testing of parenteral, ophthalmic, and topical polymyxin preparations
Key Features:
?? Higher nutrient content and pH optimized for polymyxin diffusion
?? Used exclusively with Antibiotic Medium No. 10 as the seed layer
?? Produces the clearest and largest zones for polymyxin assays
?? Globally harmonized formulation (USP/EP/JP/WHO)
Composition (per liter):
Peptone (meat or casein) � 9.0 g
Beef Extract � 2.3 g
Yeast Extract � 4.5 g
D-Glucose  � 1.5 g
Agar  � 17.0 g
Final pH: 7.2 � 0.1 at 25�C (after sterilization)
Preparation:
Suspend 34.3 g of the dehydrated medium in 1 liter of purified water. Heat with frequent agitation and boil for 1�2 minutes until completely dissolved. Sterilize by autoclaving at 121�C for 15 minutes. Cool to 60�65�C and pour 20�25 mL into each sterile 90�100 mm Petri dish to form a uniform base layer (? 5�6 mm thick). Allow to solidify on a perfectly level surface. Overlay with seeded Antibiotic Medium No. 10 containing Bordetella bronchiseptica ATCC 4617.
Storage:
Dehydrated medium: Store below 30�C in a cool, dry place.
Prepared base-layer plates: Store at 2�8�C and use within 7 days.
Mandatory combination:
Base layer = Antibiotic Medium No. 9
Seed layer = Antibiotic Medium No. 10 + Bordetella bronchiseptica ATCC 4617
For pharmaceutical quality-control and official compendial antibiotic assay only.