microbial methods for validation of brewery cip · anaerobic plate count (apc) for viable cells –...
TRANSCRIPT
Linda Bruslind
Microbial Methods for Validation of Brewery CIP
Recommendation #1: Establish a baseline
Recommendation #2: Employ HACCP
Questions to be answered first:
1. Environmental sampling or product sampling?
– Sampling: Where? How? How often? 2. Enumeration or detection?
– Identification?
Environmental Sampling
1. RODAC plate 2. Swab/sponge collection 3. Rinse water
Some basic methods employed for enumeration:
1. Standard Plate Count (SPC) or Aerobic/Anaerobic Plate Count (APC) for viable cells
– Agar plates, Petrifilm™ 2. Membrane filtration 3. ATP measurement 4. Protein measurement
Conventional Standard Plate Count (SPC)
l Plate onto suitable agar (pour or spread) – make dilutions, if necessary
SPC: Petrifilm™
l Spread 1 ml of diluent on plate that contains dried medium
l Tetrazolium added to make colonies visible
Conventional Standard Plate Count (SPC)
l Incubated under appropriate conditions for a given time
l Visible colonies counted – 25-250 colonies (statistics) – Colony Forming Units/ml
Membrane Filtration
l Filter liquid (rinse water, wort, beer) to concentrate microbes
l Place filter onto agar plate
Considerations of plating
l Appropriateness of plating medium – nutritional content – pH – use of differential and/or selective media
Plating Media
l All-purpose media (i.e. NA, PCA) l Designed for beer (i.e. UBA) l Selective/differential for beer contaminants
(i.e. Siebel media such as HLP media)
Considerations of plating
l Incubation conditions – time – temperature – oxygen exposure
Considerations of plating
l Relative number of organisms
Adenosine Triphosphate Measurement
l ATP = primary source of energy in all living cells, disappears within 2/hr after cell death
l Linear relationships between microbial ATP & bacterial numbers from 106-109 CFU/ml
l Way to measure: – Firefly luciferin –luciferase system light emitted
l Amount of light produced is directly proportional to amount of ATP added
– Can measure 102-103 fg (102-103 cells)
ATP Measurement
l Used widely for environmental sampling l Potential problem = non-microbial ATP
present l Provides rapid, on the spot monitoring of
surfaces
Protein Measurement
l Detection of surface protein residues l Immediately results l More commonly used for allergens
Microbial Identification
l Isolation of microbe l Identification technique
– Gram stain – biochemical testing – molecular technique (i.e. PCR &
sequencing)
Biofilms
l Can form on any solid surface l Difficult to remove/treat l Constant source of microbial contamination
The ASBC 2011 Winter Workshop Making the Microscope Your Friend
WHEN: Sunday January 30th, 2011 from 1:00 pm to 5:00 pm
WHERE: Widmer Brothers 929 Russell Dr. Portland, OR
CONTACT for more info and to register JESS CAUDILL-‐ [email protected], (541) 354-‐1335 or SCOTT BRUSLIND [email protected], (541) 451-‐8571