THIS DOCUMENT IS A TRANSLATION FROM A LETTER FROM IRFAQ 

10 août 2011 

Irfaq – French Institute for Applied Research in Formulation, Analysis and Quality Science

ALKAPHARM
102 avenue Gaston Roussel
Parc BIOTECH
92320 ROMAINVILLE

Villebon sur Yvette, 10 August 2011

FAO: Mr. Nicolas SALKIND

Dear Sirs,

You asked for our opinion on the foreseeable effect, in terms of the performance of the physical form
(aqueous solution or powder), of a detergent composition containing enzymes, in particular proteases
and amylases, and of the presence of cationic bactericide agents, on the performance of this
composition.

In response to your request, we would like to make the following statements:

• In all enzyme-based compositions, there is a decrease in enzyme activity over time, which is in
particular more marked in the case of proteases. This phenomenon has been traditionally
compensated for by increasing the initial enzyme concentration to ensure sufficient activity after at
least one to two years of storage at room temperature. Experts are however well aware that this
reduction of activity takes place much faster in aqueous base liquid compositions than in powders.
In the first case, it is usually necessary to add stabilising agents such as MEA borate, calcium
formiate etc. to create intramolecular bonds in the enzyme structures in order to stabilise the
position of their active sites. As for enzymes kept in solid form or immobilised on a solid substrate
(more often than not coated), it is not necessary to add these stabilising agents, as the level of
stability is three to four times higher than that of aqueous solution enzymes; enzymes in non-
aqueous polar liquid phase have a level of stability slightly higher than that measured in the
aqueous phase. However, enzyme activity is generally very inhibited in the solvent phase.
• Cationic tensioactive agents, such as quaternary ammonium derivatives traditionally used as
bactericide agents (ammonium trimethyl cetostearyl chloride, benzalkonium chloride, didecyl-
dimethyl ammonium) discourage enzyme activity by becoming fixed to the carboxyl sites of the
protein chains, blocking the active sites of the enzymes. This effect appears of course to be even
more marked if the composition is in liquid form due to the accessibility of these active sites.

To sum up, we may consider that the stability of an enzyme-based cleaning product would be
considerably affected if it were in the form of an aqueous solution, and its performance would in
particular be highly inhibited if cationic agents were added to this liquid composition.

Please do not hesitate to contact us should you have any further questions.

Yours faithfully,

[Signature]

Pierre GODDINET
CEO and Scientific Director