Brown Melanin Pigment Research Articles

Bioproduction and optimization of newly characterized melanin pigment from Streptomyces djakartensis NSS-3 with its anticancer, antimicrobial, and radioprotective properties

BackgroundMelanin is a natural pigment that is considered a promising biomaterial for numerous biotechnological applications across several industries. Melanin has biomedical applications as antimicrobial, anticancer, and antioxidant properties. Additionally, in the pharmaceutical and cosmetic industries, it is used in drug delivery and as a radioprotective agent. Also, melanin has environmental uses in the fields of bioremediation and the food industry. The biosynthesis of melanin pigment is an area of interest for researchers due to its multifunctionality, high compatibility, and biodegradability. Therefore, our present work is the first attempt to characterize and optimize the productivity of melanin pigment from Streptomyces djakartensis NSS-3 concerning its radioprotection and biological properties.ResultsForty isolates of soil actinobacteria were isolated from the Wadi Allaqui Biosphere Reserve, Egypt. Only one isolate, ACT3, produced a dark brown melanin pigment extracellularly. This isolate was identified according to phenotypic properties and molecular phylogenetic analysis as Streptomyces djakartensis NSS-3 with accession number OP912881. Plackett–Burman experimental design (PBD) and response surface methodology (RSM) using a Box-Behnken design (BBD) were performed for optimum medium and culturing conditions for maximum pigment production, resulting in a 4.19-fold improvement in melanin production (118.73 mg/10 mL). The extracted melanin pigment was purified and characterized as belonging to nitrogen-free pyomelanin based on ultraviolet–visible spectrophotometry (UV–VIS), Fourier transform infrared (FT-IR), Raman spectroscopy, scanning electron microscopy (SEM), energy dispersive X-ray spectroscopy (EDX), and NMR studies. Purified melanin demonstrated potent scavenging activity with IC50 values of 18.03 µg/mL and revealed high potency as sunscreens (in vitro SPF = 18.5). Moreover, it showed a nontoxic effect on a normal cell line (WI38), while it had a concentration-dependent anticancer effect on HCT116, HEPG, and MCF7 cell lines with IC50 = 108.9, 43.83, and 81.99 µg/mL, respectively. Also, purified melanin had a detrimental effect on the tested MDR bacterial strains, of which PA-09 and SA-04 were clearly more susceptible to melanin compared with other strains with MICs of 6.25 and 25 µg/mL, respectively.ConclusionOur results demonstrated that the newly characterized pyomelanin from Streptomyces djakartensis NSS-3 has valuable biological properties due to its potential photoprotective, antioxidant, anticancer, antimicrobial, and lack of cytotoxic activities, which open up new prospects for using this natural melanin pigment in various biotechnological applications and avoiding chemical-based drugs.

Melanin pigmentation variations in the larval cuticle of almond moth, Ephestia cautella caused by gamma radiation

To determine the effects of gamma radiation on the melanization process and phenoloxidase activity, Ephestia cautella larvae were exposed to dosages of 200, 400, 600, 800, and 1000 Gy. After irradiation, the number of non melanized larvae and the number exhibiting a slight melanization usually increased. The degree of melanization in treated larvae differed significantly from untreated larvae. The amount of melanin usually decreases as the dosage increases and as time passes after the treatment. The results of the phenoloxidase assay indicate that the enzyme activity responds differently to radiation. For instance, at doses of 200, 400, and 800 Gy, the enzyme activity remained consistent in both control and irradiated larvae. However, at doses of 600 and 1000 Gy, the enzyme activity increased to 14.92 and 13.37 O.D. units, respectively, compared to 8.81 O.D. units in the control. In order to determine if irradiated larvae have been previously exposed to ionizing radiation, a quick and easy test based on phenoloxidase activity or the melanization response is presented for use in quarantine treatment. Histological changes, specifically in the pigment granules of melanin, were studied using a light microscope. Upon inspection of the unirradiated larvae, it was observed that brown melanin pigment granules were deposited in the epicuticle and exocuticle layers of the cuticle. When gamma radiation dosages were administered to larvae, it was observed that the melanin pigment gradually diminished until it vanished at the highest dose (1000 Gy).

Camouflage of Vitiligo by Medical Tattooing.

Vitiligo is an acquired skin depigmentation disorder that affects 0.5-2% of the world population. It is characterized by loss of the natural brown melanin pigment of the skin clinically manifested as few or many white patches on the skin and microscopically with the total absence of me-lanocytes in the epidermis. The change in appearance caused by vitiligo can affect persons' emotional and psychological well-being and may cause them to alter their lifestyle. The social complication of vitiligo depends on ethnicity and on geography and local opinion, which may deem vitiligo contagious. The aim of the medical tattooing procedure in vitiligo is to revert the white vitiligo patches to normal-looking skin of natural or near natural color through installation of brownish tattoo pigment. The coloring effect is not permanent and tends to fade over time, and repeated treatment may be needed after about a year. This chapter reviews vitiligo and indications, technique, and procedures associated with medical tattooing of the disease.

Technological aspects of obtaining melanin and powder from buckwheat hull and their use in food technology

In the paper, technology of melanin and powder from buckwheat (Fagopyrum esculentum) hull obtaining is studied. Buckwheat hull is a by-product of buckwheat grain production and has high antioxidant activity. It is a cheap source for raw material with high content of cellulose and brown melanin pigment. In Russia, buckwheat grain is strategical crop used in child and dietary nutrition. Buckwheat hulls, which production volume is about 22% of the total volume of processed grain, are practically not used. At the same time, buckwheat hull can be regarded as a valuable raw material for obtaining of several biologically active substances and technological food ingredients due to its unique chemical composition. The possibility and prospects of using products of buckwheat hulls processing in the formulation of dessert with functional properties, is described. It was found that the implementation of buckwheat processing by-products into the desserts increases their antioxidant activity, and addition of fine powder increases dietary fiber content in them.

Recent advances in understanding the molecular basis of melanogenesis in melanocytes.

Melanin pigments are responsible for human skin and hair color, and they protect the body from harmful ultraviolet light. The black and brown melanin pigments are synthesized in specialized lysosome-related organelles called melanosomes in melanocytes. Mature melanosomes are transported within melanocytes and transferred to adjacent keratinocytes, which constitute the principal part of human skin. The melanosomes are then deposited inside the keratinocytes and darken the skin (a process called tanning). Owing to their dark color, melanosomes can be seen easily with an ordinary light microscope, and melanosome research dates back approximately 150 years; since then, biochemical studies aimed at isolating and purifying melanosomes have been conducted. Moreover, in the last two decades, hundreds of molecules involved in regulating melanosomal functions have been identified by analyses of the genes of coat-color mutant animals and patients with genetic diseases characterized by pigment abnormalities, such as hypopigmentation. In recent years, dynamic analyses by more precise microscopic observations have revealed specific functions of a variety of molecules involved in melanogenesis. This review article focuses on the latest findings with regard to the steps (or mechanisms) involved in melanosome formation and transport of mature melanosomes within epidermal melanocytes. Finally, we will touch on current topics in melanosome research, particularly on the "melanosome transfer" and "post-transfer" steps, and discuss future directions in pigment research.

Spinal meningeal melanocytoma in a 5-year-old child: a case report and review of literature

BackgroundMeningeal melanocytoma is considered a rare lesion arising from leptomeningeal melanocytes. Nearly two thirds of meningeal melanocytomas were reported in the intracranial compartment and the remaining one third in the spine. Spinal melanocytomas can be extradural or intradural, with extradural variant being more common, and the majority of cases have been single reports.MethodsA 5-year-old male presented with a 4-month history of non-radiating low back pain persistent at rest, with otherwise non-remarkable medical history. The patient was neurologically intact with no deficits. Preoperatively, routine laboratory investigations were non-remarkable. MRI imaging was done and showed a lesion at the level of T11 to L4, hyperintense on T1 and hypointense on T2 with homogenous contrast enhancement. Intraoperatively, the lesion was hemorrhagic, brownish, and rubbery in consistency attached to the ventral dura. Microscopic picture revealed dense cytoplasmic brown melanin pigments, with no significant mitoses or nuclear atypia. What is unique about our case is the age of the patient (5 years).ResultsTo the best of our knowledge, after reviewing the literature, this is the youngest case to be reported.ConclusionsSMM is an extremely rare tumor with a benign course. Complete surgical excision should be attempted. Age of presentation may be as young as in our case and the diagnosis of such a tumor should never be excluded in this early age group with persistent low back ache.

A catechol oxidase AcPPO from cherimoya (Annona cherimola Mill.) is localized to the Golgi apparatus

Cherimoya (Annona cherimola) is an exotic fruit with attractive organoleptic characteristics. However, it is highly perishable and susceptible to postharvest browning. In fresh fruit, browning is primarily caused by the polyphenol oxidase (PPO) enzyme catalyzing the oxidation of o-diphenols to quinones, which polymerize to form brown melanin pigment. There is no consensus in the literature regarding a specific role of PPO, and its subcellular localization in different plant species is mainly described within plastids. The present work determined the subcellular localization of a PPO protein from cherimoya (AcPPO). The obtained results revealed that the AcPPO- green fluorescent protein co-localized with a Golgi apparatus marker, and AcPPO activity was present in Golgi apparatus-enriched fractions. Likewise, transient expression assays revealed that AcPPO remained active in Golgi apparatus-enriched fractions obtained from tobacco leaves. These results suggest a putative function of AcPPO in the Golgi apparatus of cherimoya, providing new perspectives on PPO functionality in the secretory pathway, its effects on cherimoya physiology, and the evolution of this enzyme.

Polyphenol oxidase in leaves: is there any significance to the chloroplastic localization?

Polyphenol oxidase (PPO) catalyses the oxidation of monophenols and/or o-diphenols to o-quinones with the concomitant reduction of oxygen to water which results in protein complexing and the formation of brown melanin pigments. The most frequently suggested role for PPO in plants has been in defence against herbivores and pathogens, based on the physical separation of the chloroplast-localized enzyme from the vacuole-localized substrates. The o-quinone-protein complexes, formed as a consequence of cell damage, may reduce the nutritional value of the tissue and thereby reduce predation but can also participate in the formation of structural barriers against invading pathogens. However, since a sufficient level of compartmentation-based regulation could be accomplished if PPO was targeted to the cytosol, the benefit derived by some plant species in having PPO present in the chloroplast lumen remains an intriguing question. So is there more to the chloroplastic location of PPO? An interaction between PPO activity and photosynthesis has been proposed on more than one occasion but, to date, evidence either for or against direct involvement has been equivocal, and the lack of identified chloroplastic substrates remains an issue. Similarly, PPO has been suggested to have both pro- and anti-oxidant functions. Nevertheless, several independent lines of evidence suggest that PPO responds to environmental conditions and could be involved in the response of plants to abiotic stress. This review highlights our current understanding of the in vivo functions of PPO and considers the potential opportunities it presents for exploitation to increase stress tolerance in food crops.

Actinobacteria occurrence and their metabolic characteristics in the nickel-contaminated soil sample

AbstractHeavy metals are a significant source of pollution in soils that have been demonstrated to exert significant toxic effect on soil microbial assemblages. Here we investigate the occurrence and metabolic characteristics of actinobacteria, which form a predominated component of farmland bacterial community near the town of Sereď in southwest Slovakia, contaminated by close nickel ore facility. Actinobacteria occurred in this environment with high concentrations of nickel (2.109 mg/kg), slightly above the natural occurrence of cobalt (355 mg/kg) and zinc (177 mg/kg), even too low concentration of iron (35.75 mg/kg) for a normal soil and not a toxic amount of copper (32.2 mg/kg) and cadmium (<0.25 mg/kg). The phylogeny was reconstructed using partial sequences of 16S rDNA genes of both, actinobacterial isolates and clones. A total of 105 actinobacterial representatives were divided into 66 species belonging to one order, 7 genera and 5 uncategorized groups. The selected 14 morphologically distinct isolates were able to produce drop collapsing, haemolytic and lipase activities. Whereas only 4 isolates produced dark brown melanin pigment and only 5 of them produced decolourization of the azo dye, all isolates tested were capable of assimilating all 11 sugars tested. All these actinobacterial isolates were resistant to nickel, cobalt, zinc, cadmium, copper, but the level of resistance differed between the individual isolates, and the resistance profiles of antibiotics (gentamycin, ampicillin, ciprofloxacin, chloramphenicol, erythromycin, rifampicin, penicillin-G) varied among them to a certain extent. Our results suggested that actinobacteria in soil contaminated by nickel present a relatively divergent group inside of microbial assemblage.

Melanotic Xp11 Translocation Renal Cancer

Melanotic Xp11 translocation renal cancer is a recently recognized aggressive epithelioid neoplasm with features overlapping between PEComa, carcinoma, and melanoma. We describe morphologic and immunohistochemical characteristics of a melanotic Xp11 translocation renal cancer occurring in an 18-year-old girl and perform molecular genetic studies to analyze its genetic alterations and related melanogenetic activities. The tumor was composed of solid nests of epithelioid cells bearing abundant clear to finely granular eosinophilic cytoplasm and separated by delicate vascular septa. Finely granular and nonrefractile brown melanin pigments, highlighted by Fontana-Masson stain, were scattered through the tumor. By immunohistochemistry, the tumor was diffusely and strongly labeled by TFE3 and focally stained by HMB45 in a patchy pattern. In contrast, all other applied immunomarkers, including cytokeratins, epithelial membrane antigen, vimentin, CD10, S-100, smooth muscle actin, desmin, c-kit, CD68, and microphthalmia-associated transcription factor, were nonreactive to the tumor. Reverse transcription-polymerase chain reaction and validating sequencing demonstrated PSF-TFE3 gene fusion, a novel exon composition juxtaposing PSF exon 9 to TFE3 exon 5. Up-regulations of melanogenesis-associated regulators, including microphthalmia-associated transcription factor, tyrosinase (TYR), and tyrosinase-related protein 1 (TYRP1), were identified in the tumor by semiquantitative reverse transcription-polymerase chain reaction. The morphologic and immunohistochemical discrepancies between this intriguing melanotic tumor and other documented renal cell carcinomas bearing identical PSF-TFE3 gene fusion may suggest melanotic Xp11 translocation renal cancer is a distinct entity among the MiT/TFE family neoplasms.

Characterization of a Homogentisate Dioxygenase Mutant in Pseudomonas chlororaphis O6

Transposon mutagenesis of Pseudomonas chlororaphis O6 was performed with the transposon Tn5 to investigate genes involved in production of secondary metabolites. A mutant, termed Org, produced intense dark-brown pigmentation on rich medium. The Tn5-flanking sequence of the Org mutant showed high homology with the hmgA gene encoding the enzyme homogentisate dioxygenase, involved in the degradation of aromatic amino acids such as tyrosine. Growth of the hmgA mutant on L-tyrosine as sole carbon and energy sources was impaired. Growth on L-tyrosine was restored and production of the brown melanin pigment was eliminated when the mutant was complemented with the wild-type hmgA gene. The change in aromatic amino acids metabolism caused by the deletion of the hmgA gene function did not impair production of phenazines and biological traits connected to these secondary compounds: inhibition of fungal growth and inhibition of barley seed germination.

An ABC transporter from Bacillus thuringiensis is essential for beta-exotoxin I production.

beta-Exotoxin I is a nonspecific insecticidal metabolite secreted by some Bacillus thuringiensis strains. Several studies of B. thuringiensis strains that have lost the capacity to produce beta-exotoxin I have suggested that there is a strong correlation between high levels of beta-exotoxin I production and the ability to synthesize crystal proteins. In this study, we showed that a mutant strain, B. thuringiensis 407-1(Cry(-))(Pig(+)), with no crystal gene, produced considerable amounts of beta-exotoxin I together with a soluble brown melanin pigment. Therefore, beta-exotoxin I production can take place after a strain has lost the plasmids bearing the cry genes, which suggests that these curable plasmids probably contain determinants involved in the regulation of beta-exotoxin I production. Using a mini-Tn10 transposon, we constructed a library of strain 407-1(Cry(-))(Pig(+)) mutants. We screened for nonpigmented mutants with impaired beta-exotoxin I production and identified a genetic locus harboring two genes (berA and berB) essential for beta-exotoxin I production. The deduced amino acid sequence of the berA gene displayed significant similarity to the ATP-binding domains of the DRI (drug resistance and immunity) family of ATP-binding cassette (ABC) proteins involved in drug resistance and immunity to bacteriocins and lantibiotics. The berB gene encodes a protein with six putative transmembrane helices, which probably constitutes the integral membrane component of the transporter. The demonstration that berAB is required for beta-exotoxin I production and/or resistance in B. thuringiensis adds an adenine nucleotide analog to the wide range of substrates of the superfamily of ABC proteins. We suggest that berAB confers beta-exotoxin I immunity in B. thuringiensis, through active efflux of the molecule.

Biosensing of opioids using frog melanophores

Spectacular color changes of fishes, frogs and other lower vertebrates are due to the motile activities of specialized pigment containing cells. Pigment cells are interesting for biosensing purposes since they provide an easily monitored physiological phenomenon. Melanophores, containing dark brown melanin pigment granules, constitute an important class of chromatophores. Their melanin-filled pigment granules may be stimulated to undergo rapid dispersion throughout the melanophores (cells appear dark), or aggregation to the center of the melanophores (cells appear light). This simple physiological response can easily be measured in a photometer. Selected G protein coupled receptors can be functionally expressed in cultured frog melanophores. Here, we demonstrate the use of recombinant frog melanophores as a biosensor for the detection of opioids. Melanophores were transfected with the human opioid receptor 3 and used for opiate detection. The response to the opioid receptor agonist morphine and a synthetic opioid peptide was analyzed by absorbance readings in an aggregation assay. It was shown that both agonists caused aggregation of pigment granules in the melanophores, and the cells appeared lighter. The pharmacology of the expressed receptors was very similar to its mammalian counterpart, as evidenced by competitive inhibition by increasing concentrations of the opioid receptor inhibitor naloxone. Transfection of melanophores with selected receptors enables the creation of numerous melanophore biosensors, which respond selectively to certain substances. The melanophore biosensor has potential use for measurement of substances in body fluids such as saliva, blood plasma and urine.

Development of an integrated method of skin phenotype measurement using the melanins

Different types of melanin pigments have recently been identified and recognized as critical determinants of the photosensitivity of individuals. Eumelanin, the black to brown melanin pigments, are believed to protect against ultraviolet-induced cell damage, while phaeomelanin, the reddish brown variant, is thought to be photosensitizing. The relative, qualitative and absolute amount of melanin production under stimulation of solar radiation is likely to be genetically determined. The hypothesis of this study is that determination of these values can help in identifying those people who are less protected. However, these techniques must be evaluated at a population level and against traditional epidemiological measures. We assessed the amount and type of melanin in 195 subjects in four centres across Europe, relating the results to epidemiological measures such as skin characteristics, history of sunburns and number of naevi. The most important finding was that eumelanin and phaeomelanin have very different distributions in the population, being associated with other phenotype characteristics with different patterns. The relationship between phaeomelanin and eumelanin is linearly inverse in the range from black to dark blonde hair colour, while it is weakly directly proportional in the range from dark blonde to light blonde, with people with red hair showing a peculiar pattern. Phaeomelanin rather than eumelanin seemed to be independent of other skin characteristics. The results show the feasibility of a further study with an appropriate case-control design and accurate determination of melanin.

Histochemical demonstration of the activity of phosphatases by means of leadrhodizonate

A method for the histochemical demonstration of the activity of phosphatases by means of a pinkish-red staining obtained with leadrhodizonate is described. This method was specially developed for demonstrating sites of phosphatase activity in structures and cells containing melanin. The pinkish-red reaction product contrasts well with the brown melanin pigments. After drying and mounting of the sections in malinol, the staining is colour fast.