Melanin Composition Research Articles

The Structural Characteristics of Sustainable Nanomelanin Derived from Lasiodiplodia theobromae

Natural melanin is a sustainable and environmentally friendly pigment. The structural properties of melanin are of great significance for its functional applications. In this study, the preliminary composition and structure of melanin extracted from the induced cultures of Lasiodiplodia theobromae were qualitatively characterized by ultraviolet-visible (UV) spectrophotometer, elemental analysis (EA), Fourier transform infrared spectrometer (FTIR), nuclear magnetic resonance (NMR), electron paramagnetic resonance (EPR) and ultra-performance liquid chromatography-mass spectrometry (UPLC-MS). The results showed that the secreted melanin contained many functional groups such as hydroxy groups, alkanes and aromatic rings, which belonged to benzoquinone structure and aliphatic structure. The melanin secreted by Lasiodiplodia theobromae can be identified as a type of allomelanin. This research work is expected to provide a reference for the application of melanin in various fields in the future.

TECHNOLOGY FOR ISOLATION AND STUDY OF THE CHEMICAL COMPOSITION OF MELANIN FROM THE BASIDIOUS MUSHROOM BOVISTA PLUMBEA

The results of the development of a technology for the isolation of melanins from the basidiomycete mushroom Bovista plumbea collected in the territory of Tashkent and the Tashkent region of the Republic of Uzbekistan are presented. A technological scheme for obtaining melanins from basidiomycetes has been developed, which includes the following stages: drying, grinding of raw materials, purification, water extraction, filtration, concentration, acidification, centrifugation, dissolution, washing, drying. By water extraction, melanins were isolated, which are dark brown, alkali-soluble powder, with a molecular weight of 55686 Da, the content of which in the extract was 15.35%, respectively, which indicates the prospects of this type of raw material for obtaining drugs based on them. When studying the elemental composition of melanins obtained from the fruiting bodies of basidiomycetes by the method of water extraction, it was found that they contain 9.706% nitrogen, 55.831% carbon and 7.203% hydrogen. UV and IR spectroscopy was used to confirm that the obtained samples belonged to the category of melanins. The IR spectra of melanin isolated from the fruiting body of the basidiomycete were studied. Melanins are characterized by absorption bands in the region of 1690-1590 cm-1, indicating the presence of conjugated aromatic bonds in their structure. There is also a wide absorption band in the region of 3430-3100 cm-1, which is due to the presence of polyphenolic and OH groups. UV spectral analysis for the content of melanin in fractions of basidiomycete raw materials showed the presence of absorption bands in the region of 320-380 nm, corresponding to the characteristic absorption bands of melanin.

Melanin: insights into structure, analysis, and biological activities for future development.

Melanin, a widely distributed pigment found in various organisms, possesses distinct structures that can be classified into five main types: eumelanin (found in animals and plants), pheomelanin (found in animals and plants), allomelanin (found in plants), neuromelanin (found in animals), and pyomelanin (found in fungi and bacteria). In this review, we present an overview of the structure and composition of melanin, as well as the various spectroscopic identification methods that can be used, such as Fourier transform infrared (FTIR) spectroscopy, electron spin resonance (ESR) spectroscopy, and thermogravimetric analysis (TGA). We also provide a summary of the extraction methods of melanin and its diverse biological activities, including antibacterial properties, anti-radiation effects, and photothermal effects. The current state of research on natural melanin and its potential for further development is discussed. In particular, the review provides a comprehensive summary of the analysis methods used to determine melanin species, offering valuable insights and references for future research. Overall, this review aims to provide a thorough understanding of the concept and classification of melanin, its structure, physicochemical properties, and structural identification methods, as well as its various applications in the field of biology.

Histological, elemental, and ultrastructural analysis of melanin in mantle of Pacific oyster (Crassostrea gigas).

Colorful shell of bivalve is mainly because of the biological pigments, of which melanin plays an important role in shell color formation. More and more studies focus on the genes function involved in melanin synthesis, but relatively few studies address the biochemical character and ultrastructure of melanin in bivalve from microscopic perspective. Here, we investigated the histological structure of mantle of Crassostrea gigas with orange shell color. Distribution of melanin in mantle was verified with histochemical staining. In addition, immunofluorescence technique showed that strongly positive signal of CgTYR was specific to the mantle margin, which is consistence with the location of brown granules in H&E staining. The further result of elementary composition of melanin displayed that metal Ca, Fe, and Zn were detected using scanning transmission electron microscope and energy dispersive spectroscopy mapping methods. Next, based on TEM observations, it was speculated that the series of cellular events leading to the formation and release of melanin. Melanocyte in the primary stage showed many mitochondria and rough endoplasmic reticulum as well as an extensive Golgi complex with numerous vesicles intermingled with melanosome. Subsequently, melanosome was expended and their hue gradually intensified, and Golgi complex and mitochondria were still observed in the cytoplasm. Finally, after melanosome was discharged into intercellular spaces, the disintegration of membranes in some cells, and severe cellular vacuolization. These data enrich the understanding of ultrastructural characteristic and formation of melanin in mantle of bivalve and pave the way for further investigating shell coloration at the cellular level.

Effects of Aging on Hair Color, Melanosomes, and Melanin Composition in Japanese Males and Their Sex Differences.

In a previous study, we observed that the hair color of Japanese females darkens with age and that the causes of this are the increase in melanosome size, the amount of melanin, and the mol% of 5,6-dihydroxyindole (DHI) which has a high absorbance. In this study, we extended the same analyses to male hair to examine the sex differences in hair color, melanin composition, and melanosome morphology. Male hair also tended to darken with age, but it was darker than female hair in those of younger ages. Although there was no age dependence of DHI mol% in male hair, as with female hair, the melanosomes' sizes enlarged with age, the total melanin amount increased, and these findings were correlated with hair color. The analyses, considering age dependence, revealed that there were significant sex differences in the ratio of absorbance of dissolved melanin at the wavelength of 650 nm to 500 nm, in pheomelanin mol%, and in melanosome morphology parameters such as the minor axis. This may be the cause of the sex differences in hair color. Furthermore, the factors related to hair color were analyzed using all the data of the male and female hairs. The results suggested that total melanin amount, pheomelanin mol%, and DHI mol% correlated with hair color.

Characterization of Fungal Melanins from Black Stains on Paper Artefacts

Melanins play a fundamental role in the biology and ecology of several fungal species. Unfortunately, this group of amorphous macromolecules also severely (and most times irreversibly) stains cultural heritage objects. Despite efforts made throughout the years, knowledge of the chemical composition and structure of melanins is still insufficient, which hampers the task of safely cleaning these colourants from cultural heritage materials in a targeted way without causing further deterioration. This work aimed therefore to contribute towards enlightening the characteristics of fungal melanins from three fungi that are common paper colonizers: Aspergillus niger, Chaetomium globosum and Cladosporium cladosporioides. The extracted melanins were characterized by FTIR, Raman, UV-vis, Solid-State NMR and MALDI-TOF MS spectroscopies and the effect of inhibitors of DHN-melanin and DOPA-melanin pathways on colony pigmentation and growth was evaluated. Although all the extracted colourants show a predominantly aromatic structure with carbonyl and phenolic groups, some differences between the melanins can be highlighted. Melanins obtained from Ch. globosum and Cl. cladosporioides exhibited similar structures and composition and both presented DHN-melanin characteristics, while A. niger’s melanins revealed a more complex and ordered structure, with a higher prevalence of highly conjugated carbonyls than the others, besides the additional presence of a yellow/green component. These conclusions cannot be overlooked while selecting targeted cleaning methodologies for melanin stains on cultural heritage materials.

Selecting and Characterizing Tyrosinase Inhibitors from Atractylodis macrocephalae Rhizoma Based on Spectrum-Activity Relationship and Molecular Docking.

Atractylodis macrocephalae Rhizoma (AMR) is a famous classical Chinese traditional medicine (CTM), which has been used as a tonic for many diseases for thousands of years. In ancient China, it was used as a supplementary food for beauty in the palace. In preliminary studies, the function of whitening skin and the significant inhibiting effect on tyrosinase (TYR) which is the reactive enzyme in the composition of melanin of AMR were discovered, and the relevant research was rarely reported. In this study, high-performance liquid chromatography (HPLC) along with partial least squares regression analysis (PLS) was applied to survey the coherence between the chemical constituents and the inhibiting activity of 11 batches of AMR on TYR activity. The results of PLS showed that the chromatographic peaks 11 (atractylenolide III) and 15 could be important effective ingredients of the inhibition TYR activity as ascertained by spectrum-activity relationships. Furthermore, TYR inhibitory activity of atractylenolide III was validated by in vitro test by β-arbutin served as a positive control drug. The results of the in vitro test and the molecular docking showed that atractylenolide III has high TYR inhibitory activity and could link to the residues in TYR catalytic pocket. Therefore, bioassay, molecular docking, and spectrum-activity relationships are appropriate for linking the quality of samples with pharmaceutical-related active ingredients. And our studying would lay a theoretical foundation for applying the water extracts of AMR in whitening cosmetics.

Transcriptome and metabolite analysis reveals key genes for melanin synthesis during the development of Zanthoxylum bungeanum seeds

Melanin is a broad term for a family of complex organic macromolecules that contain phenol or indole groups. Melanins are found in animals, plants, and microorganisms and are widely used in medicine, food, and chemical industries. However, at present, the synthesis of plant melanin is still unclear and hinders the development and utilization of melanin. To clarify the composition of plant melanin and the key synthetic genes, we observed the cytological observation of the melanin-forming tissue of ZB seeds and analyzed the transcriptome and metabolite profiles of different developmental stages. We measured seed melanin content and gene expression levels and used redundancy analysis (RDA) to identify key genes that affected melanin synthesis. The seed melanin content was 3.85 % at the green fruit stage (S2) and reached a peak of 4.33 % at the fruit maturity stage (S4). Tyrosine and hydroquinone contents were highest at S2: 830 mg/kg and 77.23 mg/kg, respectively. Catechin, L-epicatechin, procyanidin B1, and p-coumaric acid were the main flavonoids in ZB seeds and reached their highest levels at S2. RDA analysis confirmed that S2 was the key period of melanin synthesis and that ZbPPO, ZbLAR, ZbTYRP1, and ZbTYR had crucial roles in the synthesis of ZB melanin-related substances.

Multidisciplinary characterization of melanin pigments from the black fungus Cryomyces antarcticus.

Melanin is a natural pigment present in almost all biological groups, and is composed of indolic polymers and characterized by black-brown colorization. Furthermore, it is one of the pigments produced by extremophiles including those living in the Antarctic desert, and is mainly involved in their protection from high UV radiation, desiccation, salinity and oxidation. Previous studies have shown that melanized species have an increased capability to survive high level of radiation compared with the non-melanized counterpart. Understanding the molecular composition of fungal melanin could help to understand this peculiar capability. Here, we aimed to characterize the melanin pigment extracted from the Antarctic black fungus Cryomyces antarcticus, which is a good test model for radioprotection researches, by studying its chemical properties and spectral data. Our results demonstrated that, in spite of having a specific type of melanin as the majority of fungi, the fungus possesses the ability to produce both 1,8-dihydroxynaphthalene (DHN) and L 3-4 dihydroxyphenylalanine (L-DOPA) melanins, opening interesting scenarios for the protection role against radiation. Researches on fungal melanin have a huge application in different fields, including radioprotection, bioremediation, and biomedical applications. KEY POINTS: • Isolation and characterization by multidisciplinary approaches of fungal melanins. • Discovery that pathways for producing DOPA and DHN are both active even in its extreme habitat. • Hypothesis supporting the possibility of using melanin pigment for radioprotection.

Melanin from the Lichens Cetraria islandica and Pseudevernia furfuracea: Structural Features and Physicochemical Properties.

Lichens are symbiotic photosynthesizing organisms with thalli formed by fungi and algae/cyanobacteria that possess high stress tolerance. One of the factors that contributes to the ability of a lichen to tolerate harsh environmental conditions is the presence of unique metabolites, including high-molecular-weight dark pigments termed melanins. The chemical composition and structure of lichen melanins remain poorly studied. We analyzed the elemental composition, the main functional groups, and the physicochemical properties of melanin extracted from Cetraria islandica and Pseudevernia furfuracea lichens. Based on the C/N ratio, this pigment is allomelanin. We also identified functional groups that provide photoprotective and antioxidant properties of melanin. Melanin synthesis might be an essential defense mechanism contributing to the survival of lichens under exposure to UV radiation.

Effects of Aging on Hair Color, Melanosome Morphology, and Melanin Composition in Japanese Females.

In a previous study, we showed that the size of melanosomes isolated from Japanese female hairs enlarges with age, and this affects the hair color. In this study, we analyzed the age-dependent changes in hair melanin in order to further explore the factors related to hair color changing by aging. A significant positive correlation with age was found in the total melanin amount (TM) and the mol% of 5,6-dihydroxyindole (DHI) units, while no correlation was found in pheomelanin mol%. TM showed significant correlations with hair color parameters and the melanosome volume, suggesting that hair color darkening by aging is caused by the increase in TM due to the enlargement of the size of melanosome. From the measurement of absorbance spectra on synthetic eumelanins with different ratios of DHI and 5,6-dihydroxyindole-2-carboxylic acid (DHICA), we found that the increase in DHI mol% also contributes to the darkening of hair color by aging. In addition, the level of pyrrole-2,3-dicarboxylic acid (PDCA), a marker of DHI melanin, showed a significant negative correlation with the aspect ratio of melanosome, suggesting a contribution of DHI melanin to the change in melanosome morphology by aging.

Analysis of Melanin Structure and Biochemical Composition in Conjunctival Melanocytic Lesions Using Pump-Probe Microscopy.

PurposeWe analyze melanin structure and biochemical composition in conjunctival melanocytic lesions using pump-probe microscopy to assess the potential for this method to assist in melanoma diagnosis.MethodsPump–probe microscopy interrogates transient excited-state photodynamic properties of absorbing molecules, which yields highly specific molecular information with subcellular spatial resolution. This method is applied to analyze melanin in 39 unstained, thin biopsy specimens of melanocytic conjunctival lesions. Quantitative features of the biochemical composition and structure of melanin in histopathologic specimens are assessed using a geometric representation of principal component analysis (PCA) and principles of mathematical morphology. Diagnostic power is determined using a feature selection algorithm combined with cross validation.ResultsConjunctival melanomas show higher biochemical heterogeneity and different overall biochemical composition than primary acquired melanosis of the conjunctiva (PAM) without severe atypia. The molecular signatures of PAMs with severe atypia more closely resemble melanomas than other types of PAMs. Pigment organization in the tissue becomes more disorganized as diagnosis of the lesions worsen, but nevi are more inconsistent biochemically and structurally than other lesions. Relatively high sensitivity (SE) and specificity (SP) is achieved for differentiating between various melanocytic lesions, particularly PAMs without severe atypia and melanomas (SE = 89%; SP = 87%).ConclusionsPump–probe microscopy is a powerful tool that can identify quantitative, phenotypic differences between various types of conjunctival melanocytic lesions.Translational RelevanceThis study further validates the use of pump–probe microscopy as a potential diagnostic aid for histopathologic evaluation of conjunctival melanocytic lesions.

Characterization of Molecular Composition of Bacterial Melanin Isolated from Streptomyces glaucescens Using Ultra-High-Resolution FT-ICR Mass Spectrometry

Characterization of Molecular Composition of Bacterial Melanin Isolated from Streptomyces glaucescens Using Ultra-High-Resolution FT-ICR Mass Spectrometry

Comparison of pump-probe and hyperspectral imaging in unstained histology sections of pigmented lesions.

Microscopic variations in melanin composition can be mapped through linear and nonlinear optical responses. Though instrumentation to measure linear attenuation is simple and inexpensive, the nonlinear response provides more degrees of freedom with which to spectroscopically resolve pigments. The objective of this study is to assess differences in imaging melanin contrast by comparing hyperspectral (linear) versus pump-probe (nonlinear) microscopy of unstained histology sections of pigmented lesions. The images and analysis we have presented here show that pump-probe uncovers a greater variation in pigment composition, compared with hyperspectral microscopy, and that the two methods yield complimentary biochemical information.

Raman microspectrometric study of pigments in melanized fungi from the hyperarid Atacama desert gypsum crust

The Atacama desert is amongst the oldest and driest deserts in the world, and its hyperarid core is described as ‘the most barren region imaginable’. In this study, we describe for the first time the endolithic microbial communities composed by eukaryotic microorganisms, such as melanized fungi and algae, colonizing the interior of gypsum crusts collected in the hyperarid zone of the Atacama desert. Melanin pigments present in fungi cell walls have been studied by Raman microspectroscopy. Analyses have been performed directly in the gypsum crust sustaining the colonisation of fungi and algae, on the thin sections and on the cultivated culture of the fungi from Neocatenulostroma genus. Raman spectroscopic signature of melanin has been clearly observed in all the sample types. Two broad bands located at wavenumbers 1605–1580 cm−1 and around 1350 cm−1 have been detected in all the studied samples, with slight variations in the appearance of other bands or shoulders of smaller intensity documenting either the changes in melanin composition or other biomolecules bound to melanin. Blue and green lasers (445, 514 and 532 nm) have been shown as favourable excitation for the study of melanins as the longer wavelengths induced adverse effects that mask the Raman signal. Effect of laser power level on the resulting spectra has also been discussed. Algal carotenoid pigment has been detected in isolated regions of the sample by the Raman bands of 1520, 1155 and 1002 cm−1. The strong Raman signal of fungal melanin that has been detected in the host rock from the locality characterized by extreme conditions such as Atacama desert could be considered in the discussion about feasible biomarker molecules; however, low specificity of the broad Raman signature and similarity to the spectrum of amorphous carbon must be also taken into account. Copyright © 2017 John Wiley & Sons, Ltd.

Dual-wavelength pump-probe microscopy analysis of melanin composition.

Pump-probe microscopy is an emerging technique that provides detailed chemical information of absorbers with sub-micrometer spatial resolution. Recent work has shown that the pump-probe signals from melanin in human skin cancers correlate well with clinical concern, but it has been difficult to infer the molecular origins of these differences. Here we develop a mathematical framework to describe the pump-probe dynamics of melanin in human pigmented tissue samples, which treats the ensemble of individual chromophores that make up melanin as Gaussian absorbers with bandwidth related via Frenkel excitons. Thus, observed signals result from an interplay between the spectral bandwidths of the individual underlying chromophores and spectral proximity of the pump and probe wavelengths. The model is tested using a dual-wavelength pump-probe approach and a novel signal processing method based on gnomonic projections. Results show signals can be described by a single linear transition path with different rates of progress for different individual pump-probe wavelength pairs. Moreover, the combined dual-wavelength data shows a nonlinear transition that supports our mathematical framework and the excitonic model to describe the optical properties of melanin. The novel gnomonic projection analysis can also be an attractive generic tool for analyzing mixing paths in biomolecular and analytical chemistry.

(Invited) Mechanisms of Branching Reactions in Melanin Formation - Ab Initio Quantum Chemistry Approach

Melanin, a pigment commonly found in animals, consists of two types of oligomeric unit: eumelanin (black to brown) and pheomelanin (yellow to reddish brown). The color of the skin, the hair, and the eyes is mainly determined by the ratio of eumelanin/pheomelanin production. The biosynthesis of melanin has been extensively investigated over the past 100 years. Among all reaction steps in the biosynthesis, reactions of dopachrome and dopaquinone, precursor molecules of melanin, directly affect the composition of melanin. Dopachrome is spontaneously converted into the monomer of eumelanin via or not via desorption of carboxyl group (decarboxylation). The former (decarboxylative) path is usually preferred whereas the latter (non-decarboxylative) reaction is catalyzed by metal ions such as Cu(II). Depending on the kinetic preference of the two pathways, the carboxyl moiety of eumelanin diversely changes, affecting its properties and functions. Dopaquinone can immediately undergo intramolecular cyclization, which is a reaction to form a cyclic structure, resulting in eumelanin synthesis. In a competitive manner, another reaction, thiol binding, which is a reaction to form a covalent bond with a sulfhydryl group, can prevail above a certain concentration of thiol compounds in the solution, resulting in pheomelanin synthesis. Thus, the fate of dopaquinone is directly related to the ratio of eumelanin/pheomelanin production. Mechanistic understanding of the above-mentioned reactions is a key to predict or design the properties and functions of melanin. Therefore, we make use a theoretical approach in which quantum mechanics is applied to molecular systems. Thus, we employ first principles calculation based on density functional theory. Our results show that the rate-limiting step of dopachrome conversion is deprotonation from β-carbon (β-deprotonation) at the first step. Furthermore, the subsequent steps were found to be regulated by the protonation state of the quinonoid oxygens, which are zwitterionic oxygens bound with benzene ring. Our calculations reveal that the quinonoid protonation facilitates decarboxylation. Cu(II) coordinated by quinonoid oxygens was found to protect the oxygens against protonation, leading to suppression of decarboxylation. For dopaquinone, we investigate the first step of the cyclization and the thiol binding. To evaluate the effects of substituent bound with the benzene ring, we calculate some dopaquinone-like systems. Our results show that quinones presenting their highest occupied molecular orbital (HOMO) localized at the substituent are reactive for cyclization, and that an anti-bonding shape of lowest unoccupied molecular orbital (LUMO) in the benzene-substituent region is a signature for low reactivity toward thiol compounds. This view obtained here is consistent with experiments and would provide a physico-chemical basis of melanin synthesis. Our results will be clues for designing new materials using melanin.

Chemical composition and radical scavenging activity of melanin from Auricularia auricula fruiting bodies

Melanin extracted from Auricularia auricula fruiting bodies (AAFB) was examined by element analyzer, amino acid analyzer, inductively coupled plasma-optical emission spectrometry. Elemental composition analysis revealed that main component of AAFB melanin was pheomelanin. Amino acid analysis showed that 16 amino acids were found in AAFB melanin and total amino acid content was 321. 63 mg/g. There were 13 detectable metal elements in AAFB melanin, which was rich in Ca, Fe, Cu and Zn. In addition, AAFB melanin exhibited stronger scavenging activities on 2,2-diphenyl-l-picrylhydrazyl (DPPH) radical, superoxide radical and hydroxyl radical with IC50 values of 0.18, 0.59 and 0.34 mg/mL, respectively. These results indicated that AAFB melanin might be potentially used as a natural antioxidant.

Elucidation of the chemical composition of avian melanin

Our understanding of the chemical composition of melanin remains limited, due to a paucity of direct measurements. Avian feathers have an unparalleled diversity of melanin-based color mirroring their complex chemistry. Synchrotron-based photoionization mass spectrometry is used to determine the chemical composition of melanin from samples of black, brown, grey and iridescent feathers.

Compositional changes of human hair melanin resulting from bleach treatment investigated by nanoscale secondary ion mass spectrometry

It is important to understand the influence of bleach treatment on human hair because it is one of the most important chemical treatments in hair cosmetic processes. A comparison of the elemental composition of melanin between virgin hair and bleached hair would provide important information about the structural changes of melanin. To investigate the elemental composition of melanin granules in virgin black hair and bleached hair, these hair cross-sections are analyzed by using a nanoscale secondary ion mass spectrometry (NanoSIMS). The virgin black hair and bleached hair samples were embedded in resin and smooth hair cross-sections were obtained using an ultramicrotome. NanoSIMS measurements were performed using a Cs(+) primary ion beam to detect negative secondary ions. More intensive (16) O(-) ions were detected from the melanin granules of bleached hair than from those of virgin black hair in NanoSIMS (16) O(-) ion image. In addition, it was indicated that (16) O(-) ion intensity and (16) O(-) /(12) C(14) N(-) ion intensity ratio of melanin granules in bleached hair were higher than those in virgin black hair. Nanoscale secondary ion mass spectrometry analysis of the cross-sections of virgin black hair and bleached hair indicated that the oxygen content in melanin granules was increased by bleach treatment.