Adhesion And Cohesion

Within minutes of contact with non-sterile water, the first microorganisms will adhere4. Primary colonisation is strongly influenced by the concentration of cells in the water phase. Among the spectrum of various microbial species, a clear preference for given membrane materials can be observed; one example is the preferential colonisation of polysulfone by Pseudomonas diminuta and Staphylococcus warneri as investigated by Flemming and Schaule10. In order to investigate such preferences, samples of the mixed population of a mature biofouling layer were selectively removed from the bottom (cell-membrane interface) and the top of the layer. Various membrane materials were exposed to suspensions of the two populations and the adhesion kinetics were measured. While the bottom population showed a clear preference for the material from which it was isolated, the top population did not. This makes sense as the population at the water-biofilm interface is separated from the membrane material by many layers of microorganisms and extracellular polymeric substances (EPS). Thus, manipulation of the membrane material in order to reduce microbial adhesion most probably will select for a species that adheres to that material. With time, this species will cover the surface and mask its effect. Similar observations have been made with anti-fouling coatings on ship hulls, in particular, with copper plating.

It was shown that dead cells of P. diminuta adhere at the same rate to the surface as living cells9. This indicates that these cells already carry the "glue" in suspended form, and the material which mediates adhesion as well as cohesion is the EPS. These are composed of polysaccharides, proteins, glycoproteins, lipoproteins and other macromolecules of microbial origin10. They form a slime matrix, which sticks the cells to the surface and keeps the biofilm together. Any cleaning measure has to overcome the overall binding energy of this system. This energy is not provided by covalent chemical bonds, but by weak physio-chemical interactions. In general, they can be divided into electrostatic interactions, hydrogen bonds and van der Waals interactions (fig. 1). The average binding energy ranges between 0.1-10 % of that of a covalent C-C bond, depending on the respective conformations of the macromolecules, the water content, pH value, ionic strength, temperature and other parameters. The weak binding energy of the individual bonds is increased by the fact that the EPS molecules possess many functional groups capable of interaction. If a macromolecule has 106 possible binding sites and only 10 % of them are interacting; the binding forces of the weak interactions are multiplied by a factor of 10s, resulting in a considerable stability. The mechanism by which cleaners disintegrate biofilms is based on interference with these interactions. They do contribute, but not always in the same proportion, to the overall binding forces - they vary, according to surface properties and EPS composition. Cleaners have to overcome the cohesion forces and have to address all forms of weak physio-chemical interactions.

Fig. 1: Primary adhesion of P. diminuta to potysulfone membrane material

Surfactants are a major constituent in many cleaning formulations. They will interfere with van der Waals interactions and influence the so-called hydrophobic interactions. Van der Waals interactions can be dominant in systems in which cells adhere from water to hydrophobic surfaces. Schaule" has shown that the adhesion of P. diminuta to polysulfone membranes is performed with significant participation of van der Waals interactions. This could be demonstrated by the influence of surfactants on the adhesion rate: however, in preliminary experiments, the cohesion of alginate, an extracellular polysaccharide of Pseudomonas aeruginosa, was not affected by surfactants, but was strongly influenced by electrostatic interactions2. These examples demonstrate that in different systems, different binding forces can dominate.

Phosphate, citric acid, salts, other ionic compounds and complex-formers will interfere with weak electrostatic interactions. Many of these substances are components of cleaners. It has been shown that electrostatic interactions, which are important in cohesion of EPS molecules 12, do not prevail in adhesion of P. diminuta to hydrophobic polysulfone membrane surfaces".

So-called chaotropic agents, such as urea, tetramethyl urea, guanidine hydrochloride, and others which are known from protein chemistry interfere with hydrogen bonds. They literally cause a chaos in water structure by rapidly binding water molecules, which are ripped from hydration, water surrounding proteins or polysaccharides. Hydrogen bonds represented a dominant kind of force in both adhesion and cohesion systems as described above; however, chaotropic agents are usually not constituents of cleaners. In cohesion and adhesion experiments using alginate as an EPS model, guanidine hydrochloride showed significant effects'1,12

The entanglement of the macromolecules provides an additional factor in biofilm stability. This is addressed by the use of enzymes. The problem, however, is that the EPS macromolecules are of highly variable composition and structure, and enzymes mostly are too specific to act on the entire variety. This is one reason why enzyme treatment frequently yields disappointing results in practice.

In membrane systems, it has been shown that cleaners can improve the permeability of the fouling layer without reducing the fouling layer. In an experiment, an agar gel layer was taken as a model for a hydrogel13. Even though the layer thickness is not changed by the application of the cleaner, an almost five-fold increase in the layer permeability was seen. Thus, cleaners can improve (and decrease) the permeability of the biofilm, although this is a transient effect. The results suggest that optimisation of the fouling layer is possible even if it is not removed. This, however, is not true for every given cleaner. Correct selection and tailoring of conditioning agents has been shown to significantly alter the hydraulic resistance of biofilms. Increasing the permeability is desirable (i.e. decreasing the resistance or specific resistance). Table 1 shows some examples of changes in fouling layer permeability demonstrated with model fouling layers of filter cakes from bacteria and activated sludge. In the case of formaldehyde, a commonly used disinfectant, the effects of its application is to reduce the permeability of microbial layers substantially; that is not very surprising, as formaldehyde is used as a fixation agent in microscopy. This is an experimental example with artificial model biofilms, which helps understand why the performance can deteriorate after the use of cleaners.

Table 1. Alterations in specific hydraulic resistance, rj and permeability, Lp of microbial layers due to chemical conditioning (after [13])

Table 1. Alterations in specific hydraulic resistance, rj and permeability, Lp of microbial layers due to chemical conditioning (after [13])





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