Liquid has more surfactant clean non-protein stains better

But surfactant destabilise enzyme, so liquid is expected to clean protein stains worse

The experiment proved otherwise.

Can be explained by the properties of powder:

- More builders enzyme more deactivated- Coated to protect enzyme from surfactant in powder some enzyme cannot be released as

seen by insoluble signal granules in the experiment.- Also the enzyme in liquid is protected by separating the surfactant into “structured liquid” to

reduce its inhibiting effect while storing.-1. How and which surfactants inactivate enzymes

Protease performance and stability in the wash solution can be inhibited by the presence of certain types of surfactants.

“Autolysis has been identified as the primary mode of subtilisin activity loss in a heavy-duty liquid detergent (HDLD) formulation containing 10% linear alkylbenzene sulfonate (LAS), 6% alkylether sulfate (AES), and 8% alcohol ethoxylate (AE).”(Stoner et al., 2004)

Autolysis refers to the spontaneous disintegration of the specific protein configuration of enzymes, leading to the inability to bind to stain substrates at the active sites and hence the loss of digestive activity.

The most suitable and widely used enzymes in detergents is the subtilisin class protease produced from the bacteria Bacillus licheniformis, due to its ability to exhibit acceptable levels of activity in the presence of other detergent components (Polaina & MacCabe, 2007) as well as their widespread distribution, availability and broad substrate specificity (Araújo, Cavaco Paulo, & Casal, 2008‐ ).

Subtilisin stability heavily depends on calcium binding (Bryan, 2000) (Alexander, Ruan, & Bryan, 2001).

Linear alkylbenzene sulfonates (LAS) was reported to be harmful to proteases (Kravetz & Guin, 1985) due to its ability to remove the calcium ion from the enzymes: “Anionic surfactant aggregates (micelles, lamellar structures, etc.) in the detergent formulation carry a negative charge. We hypothesize that these aggregates destabilize subtilisin by competing favorably for calcium ions with the enzymes”.(Stoner et al., 2004)

- The current trend is to formulate these liquid formulations as structured liquids with the help of salts and polymers, so that all surfactant remains in the structured liquid and enzymes remain in the aqueous phase. (Hermann et al, 1997)

- The use of granulation techniques and enzyme prilling with enzymes being encapsulated in an inert water soluble waxy substance. In powder detergents, the enzymes are mixed with the finished powders as granulates or prills. Currently, wax-coated enzyme detergent granules are being offered in colours identical to the non-coloured detergent granules. The coloured granules are termed as signal granules by the detergent manufacturers which symbolize the presence of an extra-added active ingredient in the detergent preparations. During the experiments, these signal granules were not able to dissolve even after 30

minutes with stirring some enzymes were not released.2. Builders also inactivates enzymes

- builders are the main dried component in powder detergent (about 50%), much less in liquid.

- The function of builders include softening water by removing calcium ion, to help LAS surfactant work more efficiently (LAS is intolerable to calcium?)

- Maybe LAS will attract Ca2+ too much and are occupied, hence cannot clean stains.- But builders also remove Ca2+ from the enzymes.- Thus in powder form, enzymes are more inactivated than in liquid.- Even though more builders are in powder, less surfactants are in powder too, and the

cleaning efficiency of powder is lower, both in surfactant and enzyme activities.- Singapore water is in the soft region (low soluble ion concentrations). Hence most

builders are redundant, even exert counter effect on cleaning performance.- Reason why more builders and less surfactants are packed inside powder than liquid:

economic reason. Water can be used to make up the amount of liquid detergent but cheap surfactants have to be used to do this job in solid form.

3. Why liquid detergent enzymes are less inactivated? Any stabiliser? Why not the same stabilisation in powder?

Ethoxylation is an industrial process in which ethylene oxide is added to alcohols and phenols to turn it into a surfactant. The invention of the process is attributed to Schöller and Wittwer at I.G. Farben industries. Common surfactants produced by ethoxylation include alcohol ethoxylates and alcohol ethoxysulfates.

Russell GL, Britton LN. Use of certain alcohol ethoxylates to maintain protease stability in the presence of anionic surfactants. JSurfactants Deterg 2002;5:5–10.

Alexander, P. A., Ruan, B., & Bryan, P. N. (2001). Cation-dependent stability of subtilisin. Biochemistry, 40(35), 10634-10639.

Araújo, R., Cavaco Paulo, A., & Casal, M. (2008). Strategies towards the functionalization of Subtilisin‐ E from Bacillus subtilis for wool finishing applications. Engineering in Life Sciences, 8(3), 238-249.

Bryan, P. N. (2000). Protein engineering of subtilisin. Biochimica et Biophysica Acta (BBA)-Protein Structure and Molecular Enzymology, 1543(2), 203-222.

Kravetz, L., & Guin, K. (1985). Effect of surfactant structure on stability of enzymes formulated into laundry liquids. Journal of the American Oil Chemists’ Society, 62(5), 943-949.

Polaina, J., & MacCabe, A. P. (2007). Industrial enzymes: Springer.Stoner, M. R., Dale, D. A., Gualfetti, P. J., Becker, T., Manning, M. C., Carpenter, J. F., & Randolph, T.

W. (2004). Protease autolysis in heavy-duty liquid detergent formulations: effects of thermodynamic stabilizers and protease inhibitors. Enzyme and Microbial Technology, 34(2), 114-125.

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