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      From Extraction to Advanced Analytical Methods: The Challenges of Melanin Analysis

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          Abstract

          The generic term “melanin“ describes a black pigment of biological origin, although some melanins can be brown or even yellow. The pigment is characterized as a heterogenic polymer of phenolic or indolic nature, and the classification of eu-, pheo- and allo- melanin is broadly accepted. This classification is based on the chemical composition of the monomer subunit structure of the pigment. Due to the high heterogeneity of melanins, their analytical characterization can be a challenging task. In the present work, we synthesized the current information about the analytical methods which can be applied in melanin analysis workflow, from extraction and purification to high-throughput methods, such as matrix-assisted laser desorption/ionization mass-spectrometry or pyrolysis gas chromatography. Our thorough comparative evaluation of analytical data published so far on melanin analysis has proven to be a difficult task in terms of finding equivalent results, even when the same matrix was used. Moreover, we emphasize the importance of prior knowledge of melanin types and properties in order to select a valid experimental design using analytical methods that are able to deliver reliable results and draw consistent conclusions.

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          Most cited references150

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          Melanins: Skin Pigments and Much More—Types, Structural Models, Biological Functions, and Formation Routes

          F. Solano (2014)
          This review presents a general view of all types of melanin in all types of organisms. Melanin is frequently considered just an animal cutaneous pigment and is treated separately from similar fungal or bacterial pigments. Similarities concerning the phenol precursors and common patterns in the formation routes are discussed. All melanins are formed in a first enzymatically-controlled phase, generally a phenolase, and a second phase characterized by an uncontrolled polymerization of the oxidized intermediates. In that second phase, quinones derived from phenol oxidation play a crucial role. Concerning functions, all melanins show a common feature, a protective role, but they are not merely photoprotective pigments against UV sunlight. In pathogenic microorganisms, melanization becomes a virulence factor since melanin protects microbial cells from defense mechanisms in the infected host. In turn, some melanins are formed in tissues where sunlight radiation is not a potential threat. Then, their redox, metal chelating, or free radical scavenging properties are more important than light absorption capacity. These pigments sometimes behave as a double-edged sword, and inhibition of melanogenesis is desirable in different cells. Melanin biochemistry is an active field of research from dermatological, biomedical, cosmetical, and microbiological points of view, as well as fruit technology.
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            Functions of fungal melanin beyond virulence

            Melanins are ancient biological pigments found in all kingdoms of life. In fungi, their role in microbial pathogenesis is well established; however, these complex biomolecules also confer upon fungal microorganisms the faculty to tolerate extreme environments such as the Earth’s poles, the International Space Station and places contaminated by toxic metals and ionizing radiation. A remarkable property of melanin is its capacity to interact with a wide range of electromagnetic radiation frequencies, functioning as a protecting and energy harvesting pigment. Other roles of fungal melanin include scavenging of free radical, thermo-tolerance, metal ion sequestration, cell development, and mechanical-chemical cellular strength. In this review, we explore the various functions ascribed to this biological pigment in fungi and its remarkable physicochemical properties. Functions of fungal melanin. Fungal melanins play multiple biological functions including photoprotection, energy harvest and thermoregulation by readily absorbing and transducing electromagnetic radiation. Fungal melanins also function in free radical and metal binding; protection against dehydration, chemical and mechanical stressors; and fungal development and conidiation.
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              Melanins and melanogenesis: methods, standards, protocols.

              Despite considerable advances in the past decade, melanin research still suffers from the lack of universally accepted and shared nomenclature, methodologies, and structural models. This paper stems from the joint efforts of chemists, biochemists, physicists, biologists, and physicians with recognized and consolidated expertise in the field of melanins and melanogenesis, who critically reviewed and experimentally revisited methods, standards, and protocols to provide for the first time a consensus set of recommended procedures to be adopted and shared by researchers involved in pigment cell research. The aim of the paper was to define an unprecedented frame of reference built on cutting-edge knowledge and state-of-the-art methodology, to enable reliable comparison of results among laboratories and new progress in the field based on standardized methods and shared information.
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                Author and article information

                Journal
                Int J Mol Sci
                Int J Mol Sci
                ijms
                International Journal of Molecular Sciences
                MDPI
                1422-0067
                13 August 2019
                August 2019
                : 20
                : 16
                : 3943
                Affiliations
                [1 ]MedFuture—Research Center for Advanced Medicine, “Iuliu Hațieganu” University of Medicine and Pharmacy, Louis Pasteur Street 4-6, Gh. Marinescu Street 23, 400349 Cluj-Napoca, Romania
                [2 ]Department of Pharmaceutical Analysis, Faculty of Pharmacy, “Iuliu Hațieganu” University of Medicine and Pharmacy, Louis Pasteur Street 6, 400349 Cluj-Napoca, Romania
                [3 ]Department of Pharmaceutical Botany, Faculty of Pharmacy, “Iuliu Hațieganu” University of Medicine and Pharmacy, Louis Pasteur Street 6, 400337 Cluj-Napoca, Romania
                [4 ]Department of Dermopharmacy and Cosmetics, Faculty of Pharmacy, ”Iuliu Hațieganu” University of Medicine and Pharmacy, Ion Creangă Street 12, 400010 Cluj-Napoca, Romania
                [5 ]Horia Hulubei National Institute for Physics and Nuclear Engineering, Reactorului Street 30, 077125 Măgurele, Romania
                Author notes
                [* ]Correspondence: alina.uifalean@ 123456umfcluj.ro ; Tel.: +40-752-237-387
                [†]

                These authors contributed equally to this work.

                Author information
                https://orcid.org/0000-0002-0659-8926
                https://orcid.org/0000-0001-7823-4390
                https://orcid.org/0000-0003-4098-6962
                https://orcid.org/0000-0002-6057-4386
                https://orcid.org/0000-0003-1255-9373
                https://orcid.org/0000-0003-1128-8153
                https://orcid.org/0000-0003-1297-6345
                https://orcid.org/0000-0002-0345-0993
                Article
                ijms-20-03943
                10.3390/ijms20163943
                6719904
                31412656
                6604ad0c-6281-446e-88ab-26a5708e6cf2
                © 2019 by the authors.

                Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license ( http://creativecommons.org/licenses/by/4.0/).

                History
                : 12 July 2019
                : 11 August 2019
                Categories
                Review

                Molecular biology
                melanin,eumelanin,pheomelanin,allomelanin,biopolymers,analytical methods,maldi,pyrolysis gas chromatography

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