Hi,
Generally speaking freeze-drying proteins is a risky business. Can you
clarify - where do you get the amylase from ? Is it prepared from an organic
source of some sort, or do you use recombinant enzyme (which should not have
as many problems as you list, as far as I know) ?
You can *try* to freeze dry your sample absorbed on fine silica gel - this
may give you a powder which, upon dissolution, will contain insoluble
material that might need to be filtered off.
A.G.E.
"Phil Harrison" <arsphys at cc.usu.edu> wrote in message
news:5.1.0.14.0.20030310132614.00a17e50 at cc.usu.edu...
> I have been using a preparation of alpha-amylase (Clarase) for several
> years. We use it to digest starch from plant tissues prior to
measurement
> of carbohydrates. The Clarase has a significant level of carbohydrate in
> it, giving us too high a background, so we have to remove the carbohydrate
> before we can use the enzyme prep.
>> We have been cleaning up the enzyme by dissolving it in water, dialyzing,
> then precipitating with 75% acetone. This is repeated 3-4 times, followed
> by another dialysis, then freeze drying. In the end, we get a nice,
light,
> fluffy powder. The only problems with this procedure are:
> 1. It is very time consuming
> 2. I suspect we lose some activity due to the acetone tmt.
> 3. It generates acetone which we have to redistill or dispose of.
>> Recently I tried cleaning up a test batch on a BioGel P6-DG column. I got
> excellent separation of the carbohydrates from the protein, but when I
> freeze-dried the pooled enzyme fractions, I got a gooey, syrupy mass in
the
> bottom of the flask, not a nice fluffy powder. I have seen this remedied
> by diluting the sample more before freeze drying, but that didn't seem to
> help any.
>> Now for the question (you probably thought I'd never get there!) Any
> suggestions on how to freeze-dry this sample and get a powder instead of
goo?
>> Thanks,
>> Phil
>> Phil Harrison
>> USDA-Agricultural Research Service,
> Forage and Range Research Lab
> Utah State University, UMC 6300
> Logan, UT 84322-6300
> Phone: 435-797-3209
> FAX: 435-797-3075
> e-mail: arsphys at cc.usu.edu>> ---
