## 5 Biocleaning
### Removal of Organic Materials  
Heterotrophic microorganisms (those that utilize organic substrates) are very versatile, producing a wide range of enzymes that can break down many organic materials. Treatment with enzymes such as proteases and collagenases is already used by restorers to remove organic residues from paintings, for example, without affecting the base material [111]. Although microorganisms can be the source of such specific enzymes, it can be seen that the whole cells, containing a mixture of degrading enzymes, might be a more effective treatment in some cases, as well as being more economical. They can also be effective for otherwise difficult materials, such as casein, egg yolk, oil, and animal fat [110]. For example, [PERSON] et al. [112] used cells of the bacterium _Pseudomonas stutzeri_, plus a final enzyme treatment, to remove organic residues, including animal glue residues, from wall paintings. The cost of the biological cleaning was much lower than that of conventional methods. More recently, the same group produced a new, improved, system, with an innovative carrier for _P. stutzeri_ cells, for cleaning historic wall paintings [113]. Surface contamination of culturally important buildings by traffic fumes may also be susceptible to removal by microbial cells and their enzymes [114]. Rather than using whole bacterial cells, specific degrading enzymes can be isolated from them and used where the polluting material(s) is known. The addition of these enzymes to traditional cleaning methods can also enhance the results. [PERSON] et al. [115] successfully used a mixed enzyme preparation from the bacterium _Exiguobacterium undae_, containing proteases, peptidases, nucleases, peptide ABC transporter substrate-binding proteins and a phosphatase, to remove animal glue from 3 different types of substrate, glass, stone and wood from a range of trees species (_Quercus_ sp., _Fagus sylvatica_, and _Pica abies_). Microorganisms may be used to remove graffiti paint from heritage buildings. The modified alkyl binders and other organic polymers in the paints [116] are susceptible to some polymerases and lipases in microbial cells [116]. [PERSON] et al. [117] isolated a number of paint degrading strains from a wide variety of environmental sources. The most promising bacteria belonged to the genera _Arthrobacter_, _Bacillus_, _Gordonia_, _Microbacterium_, _Pantoea_ and _Pseudomonas_, while fungi of the genus _Alternaria_ also proved interesting. In 2019, the same group tested the system on stone surfaces, using spray-painted granite and concrete coupons as the test surfaces [118]. The bacteria were grown prior to use in a culture medium enriched with powdered graffiti, to encourage the adaptation of the cells to the target paint. Macroscopic and microscopic examination, together with color and infra-red evaluations, indicated the success of the treatment. [PERSON] et al. [119; 120] examined the effectiveness of lipase enzymes extracted from various bacteria and fungi for removing blue, green, red and black felt tip marker acrylic ink from unglazed ceramic substrates. An oil-in-water emulsion alone could remove the ink, but the addition of lipase increased the efficiency. The more gentle treatment with cells or enzymes, rather than more abrasive chemical removal, is an obvious advantage for use on culturally important monuments. Microorganisms have also been suggested for removing biofilms, which contain high levels of protein, from historic buildings [121]. Biofilms are produced by microorganisms attaching to and thriving on the stone surface, and contain, besides the cells themselves, their metabolic products, especially EPS, and compounds that may be released by their lysis, such as proteins, lipids and nucleic acids [122]. In their mildest form, biofilms simply disfigure the surface, presenting a discolo red or dirty appearance; nevertheless, this requires treatment and, if possible prevention. [PERSON] et al.