M FC Agar - M FC Broth Base Rosolic Acid: Intended Use Formulae

m FC Agar • m FC Broth Base
Rosolic Acid
Intended Use Formulae
m FC Agar and m FC Broth Base are used with Rosolic Acid in Difco™ m FC Agar
cultivating and enumerating fecal coliforms by the membrane Approximate Formula* Per Liter
filter technique at elevated temperatures. Tryptose..................................................................... 10.0 g
Proteose Peptone No. 3................................................ 5.0 g
Yeast Extract................................................................ 3.0 g
Summary and Explanation Lactose...................................................................... 12.5 g
Geldreich et al.1 formulated a medium to enumerate fecal coli- Bile Salts No. 3............................................................. 1.5 g
Sodium Chloride.......................................................... 5.0 g
forms (MFC) using the membrane filter (MF) technique without Agar.......................................................................... 15.0 g
prior enrichment. Fecal coliforms (i.e., those found in the feces of Aniline Blue.................................................................. 0.1 g
warm-blooded animals) are differentiated from coliforms from Difco™ m FC Broth Base
environmental sources by their ability to grow at 44.5 ± 0.5°C.2 Consists of the same ingredients without the agar.
Many “standard methods” membrane filtration procedures Difco™ Rosolic Acid
specify m FC medium for testing water.2-4 The American Public Rosolic Acid................................................................. 1 g/vial
*Adjusted and/or supplemented as required to meet performance criteria.
Health Association (APHA) specifies m FC medium and incu-
bation at 44.5 ± 0.5°C in the fecal coliform membrane filter Directions for Preparation from
procedure, the delayed-incubation fecal coliform procedure
Dehydrated Product
and the two-layer agar method for recovering injured fecal
Difco™ m FC Agar
coliforms.2 AOAC International specifies m FC Agar for
detecting total coliforms and fecal coliforms in foods.3 1. Suspend 52 g of the powder in 1 L of purified water. Mix
thoroughly.
The U. S. Environmental Protection Agency specifies using 2. Heat with frequent agitation and boil for 1 minute to
m FC medium in fecal coliform methods for testing water by completely dissolve the powder.
the direct MF method or the delayed-incubation MF method.4,5
3. Add 10 mL of a 1% solution of Rosolic Acid in 0.2N NaOH.
Continue heating for 1 minute. DO NOT AUTOCLAVE.
Principles of the Procedure
4. If necessary, adjust to pH 7.4 with 1N HCl.
m FC Agar and m FC Broth Base contain peptones as sources
5. Test samples of the finished product for performance using
of carbon, nitrogen, vitamins and minerals.Yeast extract
stable, typical control cultures.
supplies B-complex vitamins that stimulate bacterial growth.
Lactose is a carbohydrate. Bile Salts No. 3 inhibits growth of Difco™ m FC Broth Base
gram-positive bacteria. m FC Agar contains agar as the solidi- 1. Suspend 3.7 g of the powder in 100 mL of purified water.
fying agent. The differential indicator system combines aniline 2. Add 1 mL of a 1% solution of Rosolic Acid in 0.2N NaOH.
blue and rosolic acid. 3. If necessary, adjust to pH 7.4 with 1N HCl.
Colonies of fecal coliforms are blue; non-fecal coliforms and 4. Heat to boiling. DO NOT AUTOCLAVE.
other organisms are gray to cream-colored. 5. Cool before dispensing.
6. Test samples of the finished product for performance using
stable, typical control cultures.
Difco™ Rosolic Acid
Prepare a 1% solution, dissolving 1 g in 100 mL of 0.2N NaOH.
Difco™ & BBL™ Manual, 2nd Edition

User Quality Control
Identity Specifications Cultural Response
Difco™ m FC Agar Difco™ m FC Agar or m FC Broth Base
Dehydrated Appearance: Beige with a slight blue tint to blue, free-flowing, Prepare the medium per label directions with 1% Rosolic Acid. Using the
homogeneous. membrane filter technique, inoculate and incubate plates at 44.5 ± 0.5°C
Solution: 5.2% solution, soluble in purified water upon for 24 ± 2 hours.
boiling. Without 1% Rosolic Acid: blue, very INOCULUM COLONY
slightly to slightly opalescent, may have a slight Organism ATCC™ CFU RECOVERY color
precipitate. With 1% Rosolic Acid: cranberry red,
Enterococcus 19433 103-2×103 Marked to –
slightly opalescent, may have a slight precipitate. faecalis complete
Prepared Appearance: Without 1% Rosolic Acid—Blue, slightly opalescent, inhibition
may have slight precipitate.
Escherichia coli 25922 20-80 Good Blue
With 1% Rosolic Acid—Cranberry red, slightly
opalescent, may have slight precipitate.
Reaction of 5.2% Escherichia coli
Solution at 25°C: pH 7.4 ± 0.2 (without 1% Rosolic Acid) ATCC™ 25922
Difco™ m FC Broth Base

Dehydrated Appearance: Beige with a slight blue tint to blue, free-flowing,
homogeneous.
Solution: 3.7% solution, soluble in purified water upon boil-
ing. Solution is blue, slightly opalescent, may have
a precipitate.
Prepared Appearance: Without 1% Rosolic Acid—Blue, slightly opales-
cent, may have a very fine precipitate.
With 1% Rosolic Acid—Cranberry red, slightly
opalescent, may have a slight precipitate.
Reaction of 3.7%
Solution at 25°C: pH 7.4 ± 0.2 (without 1% Rosolic Acid)
Difco Rosolic Acid
™
Dehydrated Appearance: Dark reddish-brown with metallic green particles,

free-flowing, fine crystalline powder.
Solution: 1.0% solution, soluble in 0.2N NaOH. Solution is
deep red, clear to very slightly opalescent.
Procedure Expected Results

Difco m FC Agar
™ Colonies of fecal coliforms will be various shades of blue.
1. Prepare the agar medium from the dehydrated base according Non-fecal coliforms are gray to cream-colored.
to the label directions and with the addition of the Rosolic
Acid solution. Limitation of the Procedure
2. Pour molten agar, previously cooled to 45-50°C into special A few non-fecal coliform colonies may be observed on m FC
tight-fitting plastic dishes and allow to harden. media due to the selective action of the elevated temperature
3. Roll the membrane filter used to collect the water sample and the addition of the Rosolic Acid. It may be useful to
onto the surface of the agar, so as to avoid the formation of elevate the temperature to 45 ± 0.2°C to eliminate Klebsiella
air bubbles between the filter and the agar surface. strains from the fecal coliform group.6
4. Place the dishes in plastic bags and incubate, by immersion,
in a water bath at 44.5 ± 0.2°C for 24 ± 2 hours.
References
1. Geldreich, Huff and Best. 1965. J. Am. Water Works Assoc. 57:208.
2. Eaton, Rice and Baird (ed). 2005. Standard methods for the examination of water and wastewater,
Difco™ m FC Broth 21st ed., online. American Public Health Association, Washington, D.C.
3. Horwitz (ed.). 2007. Official methods of analysis of AOAC International, 18th ed., online. AOAC
1. Prepare the broth medium from the dehydrated base International. Gaithersburg, Md.
4. U.S. Environmental Protection Agency. 1992. Manual for the certification of laboratories analyzing
according to the label directions and with the addition of the drinking water. EPA-814B-92-002. Office of Ground Water and Technical Support Division, USEPA,
Cincinnati, Ohio.
Rosolic Acid solution. 5. Bordner, Winter and Scarpino (ed.). 1978. Microbiological methods for monitoring the environment:
2. Add 2 mL of the cooled broth to sterile absorbent pads in water and wastes. Publication EPA-600/8-78-017. Environmental Monitoring and Support Laboratory,
Office of Research and Development, U.S. Environmental Protection Agency, Cincinnati, Ohio.
special tight-fitting plastic dishes. 6. Eaton, Clesceri and Greenberg (ed.). 1995. Standard methods for the examination of water and
wastewater, 19th ed. American Public Health Association, Washington, D.C.
3. Roll the membrane filter used to collect the water sample onto
the moistened absorbent pad, so as to avoid the formation
of air bubbles between the filter and the pad.
4. Place the dishes in plastic bags and incubate, by immersion,
in a water bath at 44.5 ± 0.2°C for 24 ± 2 hours.

Availability Difco™ m FC Broth Base
Difco™ m FC Agar EPA SMWW
AOAC CCAM EPA SMWW

Cat. No. 288320 Dehydrated – 100 g
288330 Dehydrated – 500 g
Cat. No. 267710 Dehydrated – 100 g
267720 Dehydrated – 500 g Difco™ Rosolic Acid
Cat. No. 232281 Vial – 6 × 1 g

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M FC Agar - M FC Broth Base Rosolic Acid: Intended Use Formulae

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m FC Agar • m FC Broth Base

Difco™ & BBL™ Manual, 2nd Edition

Difco™ m FC Broth Base

Dehydrated Appearance: Dark reddish-brown with metallic green particles,

Procedure Expected Results

Difco™ & BBL™ Manual, 2nd Edition

AOAC CCAM EPA SMWW

Difco™ & BBL™ Manual, 2nd Edition

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