Antibiotic Medium No. 19

Antibiotic Medium No. 19

Official Sporulation Medium for Bacillus Spore Preparation in Compendial Antibiotic Assays

Catalog Number: AS-1119

Overview

Antibiotic Medium No. 19 is the official standardized medium used for the preparation of spore suspensions of Bacillus subtilis ATCC 6633 and Bacillus cereus ATCC 11778 in compendial microbiological assays of antibiotics that require spore-based inoculum. It is designated as “Medium M” in USP <81>, European Pharmacopoeia 2.7.2, Japanese Pharmacopoeia (JP 17), and WHO guidelines.

The formulation contains a high concentration of manganese, a critical factor that strongly promotes sporulation. Under recommended incubation conditions, the medium consistently yields >95% phase-bright refractile spores within 5–7 days, making it indispensable for accurate and reproducible antibiotic potency testing.

Key Features & Benefits

High manganese sulfate content for maximum sporulation

Consistently produces >95% refractile spores

Internationally harmonised formulation (USP / EP / JP / WHO)

Usable as both broth (spore production) and agar (spore purity checks)

Mandatory medium for all spore-based compendial antibiotic assays

Typical Applications

Compendial Antibiotic Assays
Preparation of standardized spore suspensions for potency assays

Cylinder-plate and turbidimetric assays requiring spore inoculum

Antibiotics Commonly Assayed
Bacitracin

Tylosin

Virginiamycin

Flavophospholipol

Nourseothricin

Pharmaceutical & Feed Additive QC
Quality-control testing of pharmaceutical antibiotics

Regulatory testing of feed-grade antibiotics

Principle of the Medium

Antibiotic Medium No. 19 provides a nutrient-balanced environment enriched with manganese ions, which act as a powerful inducer of sporulation in Bacillus species. Extended incubation allows vegetative cells to fully differentiate into stable, heat-resistant spores suitable for standardized assay inoculum. The medium may also be solidified with agar to verify spore purity and morphology.

Typical Composition (per litre)

Peptone (meat or casein) – 6.0 g

Yeast Extract – 3.0 g

Beef Extract – 1.5 g

D-Glucose – 1.0 g

Manganese Sulfate (MnSO₄·H₂O) – 0.3 g (300 mg/L — critical component)

Agar (solid medium only) – 15.0–17.0 g

Final pH: 6.6 ± 0.1 at 25 °C (after sterilisation)

Preparation

Broth (Spore Production – Most Common)
Dissolve 11.8 g of dehydrated medium in 1 L purified water

Mix thoroughly until completely dissolved

Dispense into flasks or tubes

Sterilise by autoclaving at 121 °C for 15 minutes

Cool to room temperature before inoculation

Agar (Spore Purity & Maintenance)
Add 15–17 g agar per litre to the above formulation

Boil with agitation until fully dissolved

Autoclave at 121 °C for 15 minutes

Pour plates and allow to solidify

Standard Spore Production Protocol

Inoculate heavily with Bacillus subtilis ATCC 6633

Incubate at 35–37 °C with shaking for 5–7 days

Monitor microscopically until >95% phase-bright spores are observed

Harvest spores and wash 3–5 times with sterile water

Heat-shock at 65–70 °C for 30 minutes

Store purified spore suspension refrigerated

Storage & Stability

Dehydrated medium: Store below 30 °C, dry and protected from moisture

Prepared broth or agar: Store at 2–8 °C and use within 1 month

Intended Use

For pharmaceutical and feed-additive antibiotic assay only.
Mandatory medium for all spore-based compendial assays of bacitracin, tylosin, virginiamycin, and related antibiotics.
Not for clinical diagnostic, food, or veterinary use.

Quality & Compliance

Formulated to meet USP <81>, EP 2.7.2, JP 17, and WHO requirements

Manufactured under controlled conditions for batch consistency

Suitable for regulated pharmaceutical microbiology laboratories

Customs & Trade Information

HS / AHECC Code: 3821.00.00
Prepared culture media for development or maintenance of microorganisms