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Bioburden Test Problems

Bob Friedel rfriedel at perrittlab.com
Thu May 25 08:17:18 EST 2000


One more item:

As part of your validation protocol, you should add the low number of
organisms prior to the final rinse (obviously a control is included).  The
antibiotic inactivator should have contacted the filter prior to addition of
the inoculum.

Bob Friedel
Quality Assurance Manager
Perritt Laboratories
http://www.perritt.com
rfriedel at perrittlab.com

Bob Friedel <rfriedel at perrittlab.com> wrote in message
news:8gj8gq$4ft at netaxs.com...
> Antibiotics are generally inactivated enzymatically or by other chemical
> means.  Rinsing with WFI is not going to eliminate the antimicrobial
> activity.  DO NOT USE cellulose membranes as they have a history of
binding
> antibiotic materials (See below).  Then there is the potential problem of
> the antibiotic being released from the membrane upon placement into the
> broth enrichment.  The following references will give you the information
> you need:
>
> Russell, A.D., and Rogers, D.T. (1984) "Laboratory Uses of
> Antibiotic-Inactivating Enzymes", J. Antimicrob. Chemother., Vol. 14,
p.567.
>
> Corry, J.E.L., Van Doorne, H., Mossel, D.A.A. (1977) "Recovery And Revival
> Of Microbial Cells, Especially Those From Environments Containing
> Antibiotics", In: Antibiotics in Agriculture, M. Woodbine (ed), Chapter
12,
> pp.177
>
> Negretti, F. (1989) "Experimental Observations on the Bacteriological
> Controls of the Antibiotics--I. Antibacterial Activity of Membranes
Employed
> in Bacteriological Assays", J. Pharma. & Biomed. Anal.,  Vol. 7, No. 12,
p.
> 1861.
>
> Options include:
>
> a) increasing the concentration of the chemical inactivator
>
> b) using an alternate membrane filter media (e.g., polyvinylidene
difluoride
> [PVDF] or polyethersulfone [PES]), as cellulose- based membranes tend to
> bind antimicrobial materials.  If the organism(s) do not grow on the
> antibiotic-filtered membrane, there is still activity within the membrane
> (assuming that the membrane material itself was checked for antimicrobial
> activity in the absence of the antibiotic).
>
> c) increase the volume of the rinse
>
> At some point in the process, you may need to state that the material
under
> consideration has enough intrinsic antimicrobial activity to warrant the
> exclusion of the Microbial Limit Tests.  If you've conducted several
> validation trials to date (as you have indicated) without much success,
this
> data will provide supporting justification for such a decision.
>
> Also check the following website:
> http://www.cf.ac.uk/phrmy/PHRMY-STAFF/ADR.html
>
> You can e-mail Professor A.D. Russell.  He will be able to make additional
> suggestions if you can't get the references:  russelld2 at cf.ac.uk
>
> Bob Friedel
> Quality Assurance Manager
> Perritt Laboratories
> Hightstown, NJ USA
> http://www.perritt.com
> rfriedel at perrittlab.com
>
>
> R.Randolph <microbiology at usa.net> wrote in message
> news:v24qis4i11svp2tof4tk25r4t9vomi1rns at 4ax.com...
> >
> > You might consider adding SPS to your "flushings". R.Randolph
> > ==============================
> > On Fri, 31 Mar 2000 23:09:04 +1000, "Paul A. Yeatman" <pauly at eisa.net>
> > wrote:
> >
> > >I am trying to validate a number of antibiotics using a 'bioburden
test'.
> > >Due to the antimicrobial nature of the products being tested, I am
having
> > >trouble getting our test organisms to grow.
> > >
> > >The products are vancomycin and gentamycin.  Out test organisms are
> Bacillus
> > >subtilis and Pseudomonas aeruginosa (at approx 100cfu/ml).
> > >
> > >Ideally we would filter 50 mls of the product using either a 2 micron
> > >cellulose membrane, or a durapore membrane.  So far I have tried
flushing
> > >with sterile water WFI, followed by a regime of letting the membrane
sit
> > >immersed in Tryptone Soya broth  TSB for differeing amounts of time, in
> an
> > >affort to kick start the microbial growth.
> > >
> > >As it stand, I am up to about a 6000ml WFI flush, followed by 3x 20mins
> of
> > >TEB (for a total of 2000ml TEB), which is getting rather excessive.
> > >
> > >Does anyone know of a medium our media department can make that will
> > >deactivate the antimicrobial effects of these antibiotics, while
allowing
> > >our t6est organisms to grow?
> > >
> > >I have tried a method similar to what we have successfully used for a
> > >cytotoxic antimicrobial - 5500ml WFI followed by 3x 20min of TEB
(1500ml
> > >total) with a cellulose membrane, filtering only 5ml of product, but to
> no
> > >effect (returns were around 25% of the controls for gentamycin, and
worse
> > >for vancomycin).
> > >
> > >Any insight would be appecitated.
> > >
> >
>
>







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