Isolation and Characterization of Allomelanin from Pathogenic Black Knot Fungus─a Sustainable Source of Melanin

Abstract

Melanin, a widespread pigment found in many taxa, is widely recognized for its high refractive index, ultraviolet (UV) protection, radical quenching ability, metal binding, and many other unique properties. The aforementioned characteristic traits make melanin a potential candidate for biomedical, separation, structural coloration, and space applications. However, the commercially available natural (sepia) and synthetic melanin are very expensive, limiting their use in various applications. Additionally, eumelanin has been the primary focus in most of these studies. In the present study, we demonstrate that melanin can be extracted from the pathogenic black knot fungus Apiosporina morbosa with a yield of ∼10% using the acid–base extraction method. The extracted melanin shows irregular morphology. Chemical characterization using X-ray photoelectron spectroscopy, infrared spectroscopy, and solid-state nuclear magnetic resonance spectroscopy reveals that the melanin derived from black knots is the less explored nitrogen-free allomelanin. Additionally, the extracted melanin shows broadband UV absorption typical of other types of melanin. Because of the wide availability and low cost of black knots and the invasive nature of the fungus, black knots can serve as an alternative green source for obtaining allomelanin at a low cost, which could stimulate its use as an UV light absorber and antioxidant in cosmetics and packaging industries.

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

“…but the pigment was found to be irregular in shape size by SEM, and it is likely this was probably destroyed by repeated acid–base extraction. Similarly, Singla et al 62 observed that allomelanin extracted from pathogenic black knot fungus was disrupted and showed irregular morphology, and it was uncertain whether this structure was present in the pathogenic bacteria. The extraction method of neuromelanin is complicated because it requires the isolation of melanin granules from brain cells.…”

“…62 Allomelanin from pathogenic black knot fungus has two absorption peaks: the peak at 1381 cm À1 is associated with the aromatic C-C linear stretching or C-N stretching of the indole structure, while the peak at 1593 cm À1 is attributed to the CQC bond of the indole structure. 62 The allomelanin of C. islandica had a distinct peak in the range of 1020 cm À1 associated with the stretching vibration of the (C-O-C) group; no absorption was recorded at 1720 cm À1 , which may indicate the absence of free carboxyl groups. 61 Pyomelanin was found with a broad absorption in the 3300-3420 cm À1 range, which corresponds to the stretching vibrations of -NH and -OH.…”

“…By light and X-ray scattering studies, McCallum et al 44 determined the dispersion, morphology and fine structure of allomelanin in solution. Singla et al 62 determined by XPS that the nitrogen content of allomelanin extracted from pathogenic black knot fungus was very low (3.5 AE 0.9%).…”

“…87 Therefore, subsequent observations by SEM magnification revealed that pheomelanin are spherical nanoparticles with an unusual size distribution, with diameters of about 90–180 nm. 54 Natural allomelanin extracted from pathogenic black knotted bacteria was irregularly shaped, 62 which may result from disruption after passing through acidic and alkaline solutions. To better understand the structure of allomelanin, Zhou et al 65 synthesized artificial allomelanin and observed that the dimer could be formed by forming an oval platelet shape and finally assumed a spherical particle shape.…”

Interactions of Melanin with Electromagnetic Radiation: From Fundamentals to Applications

“…The alkali hydrolytic treatment, on the other hand, is based on the soluble nature of melanin in the alkali. The melanized tissue is first solubilized in the alkali, followed by melanin precipitation using acid, which is normally repeated several times for complete melanin precipitation. , This method results in irregular melanin aggregates that can be contaminated by sulfur from the hydrolysis of sulfur-containing amino acids, particularly for those keratinous sources (e.g., hairs) . On the other hand, enzymatic digestion method is a milder alternative that produces intact melanin granules that are morphologically and chemically unchanged .…”