Lysozyme reaction

Lysozyme reaction

Post by Brian Lundee » Wed, 13 Dec 2000 03:54:59



I got some Lysozyme for preventing ML and yesterday I put some in a carboy
of Pinot Noir whose biological stability I'm not too sure about. It's about
2 years old and ready to bottle but I have never tested for completion of
ML, for which it was innoculated towards the end of primary fermentation. It
never showed any outward signs of going through ML, so either the innoculum
didn't take, or it was just darn sneaky in going about its business. As I
said, it's two years old, but I know if it is going to take off again, it
will do so after bottling.

The instructions say to use from 25-50 grams/hectoliter for stabilizing the
wine, so I put in one teaspoon, which if it is anything like other dry
materials I use is about 5-6 grams. For 23 liters, that puts me on the lower
end of the recommended range. I mixed it into a cup of wine, along with a
1/4 tsp of potassium metabisulfite (my bottling dose), and the "solution"
(if the word dissolve even applies here) turned a nice murky pink, kind of
like milk with a little red food coloring. When I stirred it into the
carboy, the murkiness immediately took hold in there, and then flocculation
began. The carboy was full of little tiny clumps, and when I gave the carboy
a twist, they formed a little tail and started floating upward! Eventually,
they must have started obeying gravity (not just a good idea, it's the law)
and this morning I have about a one inch layer of white sediment at the
bottom. The wine has cleared sufficiently again that I can read my light
bulb on the other side, although there are still some clumps in suspension
that are affecting the clarity. Hopefully these will all settle out, as my
wine was brilliantly clear before doing this.

The question is, what happened? The literature makes no mention of this kind
of thing happening. It does say that a haze can form when used in
conjunction with metatartaric acid. What the heck is metatartaric acid? Is
that the same as the tartaric acid I buy from the supply shop? I have added
some of that in the past (not with the lysozyme), but what I'm seeing
certainly goes behind my usual definition of a haze. BTW, the wine has never
had any fining done to it. Perhaps it attracted some proteins and/or tannins
to itself, and caused them to flocculate and settle out. Once it clears a
bit more, I am going to have to sample it and see if this has had any
negative effects on the taste and aroma.

Any thoughts?

Brian

 
 
 

Lysozyme reaction

Post by jackkelle » Wed, 13 Dec 2000 12:23:40


Brian, I have never used Lysozyme but have heard it was a good
product.  It is a protein that catalyzes the hydrolysis of certain
glucosidic linkages occurring in the cell walls of microorganisms.  It
is widely distributed in animals and plants, but enzymatic Lysozyme is
made from chicken egg whites.  Having said all of that, I cannot really
say what happened, but it undoubtedly reacted with the ML culture in
the wine and you saw the result.

Why not contact the producer of the product you used and ask...?

Jack Keller, The Winemaking Home Page,
http://winemaking.jackkeller.net/index.asp

Sent via Deja.com http://www.deja.com/
Before you buy.

 
 
 

Lysozyme reaction

Post by Charlie » Wed, 13 Dec 2000 12:40:44


Well..let's see...

your intention was to destroy any MLF bacteria still lurking in your wine,
right...the lysozymes attack and destroy the peptidoglycan layer in their
cell walls.  I think egg white fining will do this also .. same enzyme.

I just skimmed through my notes from a Food Colorants course, recalling that
some enzymes are implicated in anthocyanin rxn's and polymerization.  I only
find mention of glycosidase, peroxidase, and polyphenol oxidase.  The best
sign that one of these guys is at work would be some sort of color change
(did you notice an overall color change?).. It might be possible that the
lysozyme took on some MLF bacteria (or other MO's  :^) in your wine .. but I
doubt it.  Judging by the amount of precipitate you generated, I would say
you just "fined" your wine, whether you wanted to or not .. and probably
took out some compounds you wanted to keep, too boot!  Do you know anything
else about the stuff you put in?   What else might have been in the mix?
Also ... what did the wine "taste" like ... before "and" after (I know you
mentioned that  ...did you do it yet?)???

Charlie


Quote:
> I got some Lysozyme for preventing ML and yesterday I put some in a carboy
> of Pinot Noir whose biological stability I'm not too sure about. It's
about
> 2 years old and ready to bottle but I have never tested for completion of
> ML, for which it was innoculated towards the end of primary fermentation.
It
> never showed any outward signs of going through ML, so either the
innoculum
> didn't take, or it was just darn sneaky in going about its business. As I
> said, it's two years old, but I know if it is going to take off again, it
> will do so after bottling.

> The instructions say to use from 25-50 grams/hectoliter for stabilizing
the
> wine, so I put in one teaspoon, which if it is anything like other dry
> materials I use is about 5-6 grams. For 23 liters, that puts me on the low
er
> end of the recommended range. I mixed it into a cup of wine, along with a
> 1/4 tsp of potassium metabisulfite (my bottling dose), and the "solution"
> (if the word dissolve even applies here) turned a nice murky pink, kind of
> like milk with a little red food coloring. When I stirred it into the
> carboy, the murkiness immediately took hold in there, and then
flocculation
> began. The carboy was full of little tiny clumps, and when I gave the
carboy
> a twist, they formed a little tail and started floating upward!
Eventually,
> they must have started obeying gravity (not just a good idea, it's the
law)
> and this morning I have about a one inch layer of white sediment at the
> bottom. The wine has cleared sufficiently again that I can read my light
> bulb on the other side, although there are still some clumps in suspension
> that are affecting the clarity. Hopefully these will all settle out, as my
> wine was brilliantly clear before doing this.

> The question is, what happened? The literature makes no mention of this
kind
> of thing happening. It does say that a haze can form when used in
> conjunction with metatartaric acid. What the heck is metatartaric acid? Is
> that the same as the tartaric acid I buy from the supply shop? I have
added
> some of that in the past (not with the lysozyme), but what I'm seeing
> certainly goes behind my usual definition of a haze. BTW, the wine has
never
> had any fining done to it. Perhaps it attracted some proteins and/or
tannins
> to itself, and caused them to flocculate and settle out. Once it clears a
> bit more, I am going to have to sample it and see if this has had any
> negative effects on the taste and aroma.

> Any thoughts?

> Brian

 
 
 

Lysozyme reaction

Post by Jack Web » Wed, 13 Dec 2000 12:57:32


Jack, can you put this into laymans language?  Your 10$ words
are beyond my budget.


Quote:
> Brian, I have never used Lysozyme but have heard it was a good
> product.  It is a protein that catalyzes the hydrolysis of certain
> glucosidic linkages occurring in the cell walls of microorganisms.  It
> is widely distributed in animals and plants, but enzymatic Lysozyme is
> made from chicken egg whites.  Having said all of that, I cannot really
> say what happened, but it undoubtedly reacted with the ML culture in
> the wine and you saw the result.

> Why not contact the producer of the product you used and ask...?

> Jack Keller, The Winemaking Home Page,
> http://winemaking.jackkeller.net/index.asp

> Sent via Deja.com http://www.deja.com/
> Before you buy.

 
 
 

Lysozyme reaction

Post by jackkelle » Wed, 13 Dec 2000 22:42:48


Quote:
> Jack, can you put this into laymans language?  Your 10$ words
> are beyond my budget.

I should simply bow to Charlie G.'s explanation, but you asked.

Firstly, Lysozyme is a protein.  This is important to know should you
need to fine the wine to clear traces of the Lysozyme.  Just use an
anti-protein fining agent.

Secondly, it works by attacking glucosides (more correctly, it promotes
the chemical breakdown of glucosides, effectively eroding the cell
walls) in the cell walls of microorganisms.  Obviously, it works fast.

Jack Keller, The Winemaking Home Page,
http://winemaking.jackkeller.net/index.asp

Sent via Deja.com http://www.deja.com/
Before you buy.

 
 
 

Lysozyme reaction

Post by Lum » Thu, 14 Dec 2000 01:30:42



Quote:
>Snip..... end of the recommended range. I mixed it into a cup of wine, along
with a
> 1/4 tsp of potassium metabisulfite (my bottling dose), and the "solution"
> (if the word dissolve even applies here) turned a nice murky pink, kind

of.....snip

Hi Brian,

I have never used lysozyme.  However, adding sulfite to a small quantity of
water produces a low pH solution.  Perhaps the lysozyme is sensitive to pH (or
perhaps sulfite is sensitive to lysozyme).

BTW, the general practice is to add fining materials one at a time.  Mixing two
or more materials often causes trouble.  Of course, there are several
exceptions.

I can assure everyone that fining red wines with egg whites will _not_ prevent
MLF.

Regards,
lum

 
 
 

Lysozyme reaction

Post by Brian Lundee » Thu, 14 Dec 2000 01:36:27



Quote:
> Why not contact the producer of the product you used and ask...?

Well, that would have been a little too obvious a route to take for me to
think of. ;-)

I will let you know how it turns out.

Brian

 
 
 

Lysozyme reaction

Post by Charlie » Thu, 14 Dec 2000 02:23:42


Hi Lum ... my biochem text clearly states that egg whites are a source of
lysozyme (as well as human tears :^P  ), so I would have thought egg white
fining would have knocked out any remaining MO's ... but it didn't say "how
much" lysozyme is in egg whites .. maybe not enough!

I don't have any experience with it, tho (I don't fine OR filter my Pinot
noirs) so I will yield the floor...

Thanks,
Charlie


Quote:

> I can assure everyone that fining red wines with egg whites will _not_
prevent
> MLF.

> Regards,
> lum

 
 
 

Lysozyme reaction

Post by Allan Doble » Thu, 14 Dec 2000 03:34:09


So I suppose crying in your wine would work too?
Quote:

> Hi Lum ... my biochem text clearly states that egg whites are a source of
> lysozyme (as well as human tears :^P  ), so I would have thought egg white
> fining would have knocked out any remaining MO's ... but it didn't say "how
> much" lysozyme is in egg whites .. maybe not enough!

> I don't have any experience with it, tho (I don't fine OR filter my Pinot
> noirs) so I will yield the floor...

> Thanks,
> Charlie



> > I can assure everyone that fining red wines with egg whites will _not_
> prevent
> > MLF.

> > Regards,
> > lum

 
 
 

Lysozyme reaction

Post by Brian Lundee » Thu, 14 Dec 2000 04:48:52



Quote:
> I have never used lysozyme.  However, adding sulfite to a small quantity
of
> water produces a low pH solution.  Perhaps the lysozyme is sensitive to pH
(or
> perhaps sulfite is sensitive to lysozyme).

I spoke with a rep from Inovatech (the manufacturer) in Abbotsford, BC, a
very helpful woman named Jennifer. She did mention that I should have
dissolved the lysozyme (their brand name is Inovapure) in water (10%
solution) then added it in. It seems I didn't get the all important "how to
use this stuff" documentation, but Jennifer will be faxing that to me. She
didn't think adding the potassium metabisulfite with it would cause any
problems. In any event, I have another wine (a plonky white that most likely
will be good only for distillation) that I am going to try a lysozyme
treatment on and see how it reacts. This won't necessarily advance my
understanding of what happened in the Pinot Noir, but I'm curious to see
what happens anyway. Once I get some results back from Inovatech, I guess
the ultimate test will be to try another addition to my Pinot carboys, and
see what happens.

In any case, Jennifer has graciously offered to analyze a treated and
untreated sample of my Pinot Noir to see if it has a bacterial infection
(either lactic producing or Pediococcus,which is also blitzed by lysozyme).
In fact, Inovatech has a local office here in Winnipeg that I can drop off
the samples with, and they will send it out to their BC branch. She also
thought that their product might react with some phenolics and cause some to
drop out. However, it's also possible this is nothing more than improperly
adding the product into the wine.

I'm certainly pleased with the support I'm getting from Inovatech on this
matter, and just want to send out a public thanks to them for it.

Brian

 
 
 

Lysozyme reaction

Post by jackkelle » Thu, 14 Dec 2000 10:42:23


Brian, that's good to hear.  I'm a big believer in turning to the
manufacturer for answers first, but don't really mind seeing this
discussion.

By the way, I agree with Lum that egg whites will not stop MLF.
Lysozyme may be derived from egg whites, but the important word is
"derived."  Quite a few cosmetics are derived from coal, but rubbing
coal-dust on your face will not achieve the same results.

Jack Keller, The Winemaking Home Page,
http://winemaking.jackkeller.net/index.asp

Sent via Deja.com
http://www.deja.com/

 
 
 

Lysozyme reaction

Post by Lum » Thu, 14 Dec 2000 10:52:09


Thanks for the info Brian.  Sounds like pretty good service from Inovatech.
Please keep us informed.
Regards,
lum


Quote:
> I spoke with a rep from Inovatech (the manufacturer) in Abbotsford, BC, a
> very helpful woman named Jennifer. She did mention that I should have
> dissolved the lysozyme (their brand name is Inovapure) in water (10%
> solution) then added it in. It seems I didn't get the all important "how to
> use this stuff" documentation, but Jennifer will be faxing that to me. She
> didn't think adding the potassium metabisulfite with it would cause any
> problems. In any event, I have another wine (a plonky white that most likely
> will be good only for distillation) that I am going to try a lysozyme
> treatment on and see how it reacts. This won't necessarily advance my
> understanding of what happened in the Pinot Noir, but I'm curious to see
> what happens anyway. Once I get some results back from Inovatech, I guess
> the ultimate test will be to try another addition to my Pinot carboys, and
> see what happens.

> In any case, Jennifer has graciously offered to analyze a treated and
> untreated sample of my Pinot Noir to see if it has a bacterial infection
> (either lactic producing or Pediococcus,which is also blitzed by lysozyme).
> In fact, Inovatech has a local office here in Winnipeg that I can drop off
> the samples with, and they will send it out to their BC branch. She also
> thought that their product might react with some phenolics and cause some to
> drop out. However, it's also possible this is nothing more than improperly
> adding the product into the wine.

> I'm certainly pleased with the support I'm getting from Inovatech on this
> matter, and just want to send out a public thanks to them for it.

> Brian

 
 
 

Lysozyme reaction

Post by Charlie » Thu, 14 Dec 2000 12:38:48


Jack and Lum,

Can one of you please expand on your position that egg whites will "not"
stop MLF?  How have you determined this?   Paper chromatography after
fining? (good, but not terribly accurate, as somebody pointed out in this ng
a few weeks ago) ..   HPLC or MS(much more accurate!) ... I'm just curious,
because the way I understand it, lysozyme is an enzyme not "derived" from
egg whites, but "found" in egg whites.

I must reiterate, this is just curiosity on my part .. as I said, I've never
"fined" a wine (in fact, I've never tried to "stop" MLF either), but it sure
seems to me that what happened when the original poster added "lysozyme" was
"fining" or protein stabilization, and I, for one, am still curious if it
arrested ML as well!

Regards,
Charlie


Quote:
> By the way, I agree with Lum that egg whites will not stop MLF.
> Lysozyme may be derived from egg whites, but the important word is
> "derived."  Quite a few cosmetics are derived from coal, but rubbing
> coal-dust on your face will not achieve the same results.

> Jack Keller, The Winemaking Home Page,
> http://winemaking.jackkeller.net/index.asp

> Sent via Deja.com
> http://www.deja.com/

 
 
 

Lysozyme reaction

Post by jackkelle » Fri, 15 Dec 2000 09:22:37


Charlie, it has been too long since I read about this, but my
understanding is that enzymatic lysozyme is indeed "found" in egg
white, but it is bound inside cells of proteinous matter and must be
extracted.  This is not a monumental feat, but it isn't something you
can do with your home chemistry set either.  While naturally bound in
the egg white, it is not readily usable for other uses (of which there
are many).  In any case, the amount found in an average chicken egg is
grossly insufficient to stop MLF even if it were readily usable for
that purpose.  I don't recall how many eggs yield an effective dose,
but it seemed to me it was quite a few.  So, given that it is bound and
not very useful until extracted (as a granular powder), and that it
requires more eggs to yield an effective dose than one would use in
fining with egg white, it is fairly safe to say that fining with egg
whites will not stop MLF.  However, I find all of this rather academic
since it is VERY easy and far cheaper to stop MLF with potassium
metabisulfite.

Jack Keller, The Winemaking Home Page,
http://winemaking.jackkeller.net/index.asp

Sent via Deja.com
http://www.deja.com/

 
 
 

Lysozyme reaction

Post by Charlie » Sat, 16 Dec 2000 13:41:31


Hi Jack .. this sounds odd to me, that an enzyme whose role in nature is to
destroy bacterial cell walls, is found "bound" up inside of cells of
"proteinous matter", _unavailable_ to do it's job! ... why would egg white
cells accumulate the darn things if they can't deploy them on something as
simple as MLF bacteria?

Oh well, I agree ... academic - this thread started with a discussion about
using a lysozyme to "stabilize MLF", and causing instead (or in _addition_
to ... my point) fining!

I'm still waiting to hear whether Brian, indeed, arrested MLF _in addition_
to fining his Pinot Noir (or whether there was any malic there to begin with
:^)

Charlie


Quote:
> Charlie, it has been too long since I read about this, but my
> understanding is that enzymatic lysozyme is indeed "found" in egg
> white, but it is bound inside cells of proteinous matter and must be
> extracted.  This is not a monumental feat, but it isn't something you
> can do with your home chemistry set either.  While naturally bound in
> the egg white, it is not readily usable for other uses (of which there
> are many).  In any case, the amount found in an average chicken egg is
> grossly insufficient to stop MLF even if it were readily usable for
> that purpose.  I don't recall how many eggs yield an effective dose,
> but it seemed to me it was quite a few.  So, given that it is bound and
> not very useful until extracted (as a granular powder), and that it
> requires more eggs to yield an effective dose than one would use in
> fining with egg white, it is fairly safe to say that fining with egg
> whites will not stop MLF.  However, I find all of this rather academic
> since it is VERY easy and far cheaper to stop MLF with potassium
> metabisulfite.

> Jack Keller, The Winemaking Home Page,
> http://winemaking.jackkeller.net/index.asp

> Sent via Deja.com
> http://www.deja.com/