Isolation and characterization of melanin from black garlic and sepia ink
Introduction
Melanins are dark brown to black colored biomacromolecules formed by oxidative polymerization of phenolic or indolic compounds (Wakamatsu & Ito, 2002), which are commonly found in animals, plants, and microorganisms (Butler & Day, 1998; Kollosa, 1991). Melanins are hydrophobic and negatively charged macromolecule that is insoluble in water, acids, or organic solvents (Langfelder, Streibel, Jahn, Haase, & Brakhage, 2003). Biological functions of melanins have been well recognized even though their chemical structure has not been fully identified since they are complex and varied depending upon the combination of monomeric units during polymerization. Melanins play important roles in microorganisms against thermal, chemical, and biochemical stresses (Allam & El-Zaher, 2012), in plants as a strengthener for the cell wall (Riley, 1997), and in humans for determining the skin color and protecting against UV radiation (Huang & Chang, 2012). Functions of melanins such as ultraviolet absorbers, cation exchangers, drug carriers, amorphous semiconductors, X-ray absorbers, and contrast agent for imaging (Ju et al., 2013; Ye et al., 2014) have been used in cosmetics, functional foods, solid plastic films, lenses, paints, and other surface protection applications (Kalka, Mukhtar, Turowski-Wanke, & Merk, 2000). Melanins also have important biological functions such as antimicrobial, antitumor, antivenin, anti-inflammatory, and liver protecting functions (El-Obeid, Al-Harbi, Al-Jomah, & Hassib, 2006; Kurian, Nair, & Bhat, 2015; Manivasagan, Venkatesan, Senthilkumar, Sivakumar, & Kim, 2013; Sava, Hung, Blagodarsky, Hong, & Huang, 2003).
Biological melanins are divided into eumelanin, pheomelanin, and allomelanin based on their monomer units and synthesis process. Eumelanin is the predominant melanin synthesized in animals and microorganisms that produced by oxidative polymerization of tyrosine or phenylalanine to L-3,4-dihydroxyphenylalanine (l-DOPA) which further converted into dopachrome and then to melanin (Langfelder et al., 2003; Tarangini & Mishra, 2013). Pheomelanin is yellow to red containing sulfur formed through cysteinylation (Nappi & Ottaviani, 2000). Allomelanin is commonly produced in plant and formed from a variety of sources like dihydrofolate, homogentisic acid, catechols, etc. (Plonka & Grabacka, 2006; Varga, Berkesic, Darulad, Maye, & Palágyi, 2016). Melanins have been obtained from various sources including bacteria (Aghajanyan et al., 2005; Yuan, Burleigh, & Dawson, 2007), fungi (Langfelder et al., 2003), catfish (Kumar & Joy, 2015), cuttlefish (Mbonyiryivuze et al., 2015b; Mbonyiryivuze, Mwakikunga, Dhlamini, & Maaza, 2015a), chestnut shell (Yao & Qi, 2016), and black oat (Varga et al., 2016). As a potential source of new melanin, we focused on black garlic. Black garlic is black colored garlic browned by the Maillard reaction through aging of the whole bulb of garlic under specific conditions of temperature and humidity for 2–3 months (Kimura et al., 2017). There are many reports on the functional materials and beneficial health effect of black garlic (Angeles, Jesus, Rafael, & Tania, 2016; Kim, Kang, & Gweon, 2013; Kimura et al., 2017; Lu, Li, Qiao, Qiu, & Liu, 2017). However, no research has been reported on the black garlic pigment.
In the present study, the black pigment of black garlic was extracted using alkali treatment and characterized, and compared with representative eumelanin, sepia melanin.
Section snippets
Materials
Black garlic was obtained from a local manufacturing company. Food-grade sepia ink paste (sepia ink 92%, NaCl 8%) was purchased from Cerezo Berzosa, S. A. (Carcer, Spain). Reagent grade sodium hydroxide, ethyl alcohol (anhydrous, 99.9%), and hydrogen chloride were purchased from Daejung Chemicals & Metals Co., Ltd (Siheung, Gyeonggi-do, Korea). Gallic acid and 2,2-Diphenyl-1-picrylhydrazyl (DPPH) were procured from Sigma-Aldrich (St. Louis, MO, USA).
Isolation of melanin
Black garlic was ground first to make a paste
Morphology
The SEM micrographs of melanin isolated from black garlic (BG) and sepia ink (SI) are shown in Fig. 1. At a scale of 2 μm and 20.0 K X magnification, BG melanin appeared as aggregates with spherical granules with different sizes in the range of 100–1200 nm. On the contrary, the SI melanin observed with same magnification appeared to be aggregated with different sizes that were agglomerated together. These aggregates were also formed with small spherical granules with different sizes in the
Conclusions
Melanins were isolated from black garlic and sepia ink. Both melanins exhibited similar physicochemical properties with high UV-barrier, thermal stability, and antioxidant activity. Between the two types of melanins, SI melanin was less in size and showed higher UV-barrier and thermal stability. Both BG and SI melanin showed characteristic FTIR spectra of melanin with the existence of functional groups (CH, COOH, and NH) responsible for the binding sites with various functional groups such as
Acknowledgment
This research was supported by the Agricultural Research Council (ARC 710003) program of the Ministry of Agriculture, Food and Rural Affairs, Republic of Korea.
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