Sepia Officinalis Ink Research Articles

Natural melanin nanoparticles (MNPs) extracted from Sepia officinalis: A cost-effective, chemo-photothermal, synergistic nanoplatform for osteosarcoma treatment

Osteosarcoma conventional chemotherapeutics are known for their side effects, limited options, and induction of drug resistance. This creates the need to develop new therapeutics capable of effectively destroying cancer cells with low toxicity, improving patient survival rate and their life quality. This work reports a novel drug delivery nanoplataform made of Natural Melanin Nanoparticles (MNPs), obtained from Sepia officinalis ink, with 99% incorporation efficiency of doxorubicin (Dox) without the use of non-toxic solvents. A significant photothermal effect was shown by a 36ºC increment after 10 min of laser irradiation, surpassing reported values for synthetic melanin. A sustained drug release of ca. 23% with photothermal stimuli was observed, compared to 15% without stimuli, after 48 h. This nanoplatform is obtained as a food industry side product, which makes it a natural cost-effective biomedical material. Natural MPs were applied in an osteosarcoma cell line (SaOs-2), and internalized by the cells in less than 2 h, showing cytocompatibility up to 1000 µg/mL after 72 h of contact with cells. On the contrary, when natural MNPs loaded with Dox (Dox-MNPs) were placed in contact with the SaOs-2 cells and were simultaneously receiving NIR light it was observed a 93% reduction in cancer cells in 48 h, revealing a synergistic effect between chemotherapy and phototherapy. To our knowledge this is the first time that natural MNPs extracted from Sepia officinalis were tested on an osteosarcoma cell line as chemo-photothermal agent, showing these NPs are an effective, cost-effective, reproducible, non-toxic nanoplatform for osteosarcoma treatment using combined effects.

Sepia officinalis ink mitigates gastric ulcer via modulation of antioxidant/anti-inflammatory pathways

BackgroundSepia officinalis ink is a bioactive secondary metabolite rich in melanin granules, which has a wide range of nutritional and therapeutic values and also has been used to prevent various gastrointestinal disorders. Gastric ulcer, the most common gastrointestinal disease, is characterized by severe gastric mucosa damage, and its prevention is currently one of the main goals of clinical and experimental studies. Thus, the present study was focused on evaluating the potential gastroprotective efficacy of Sepia officinalis ink extract (SOIE) against ethanol-induced gastric ulcer in rats.ResultsThe current results revealed that SOIE administration at the two selected doses improved significantly gastric mucosa integrity as indicated by the significant (P < 0.05) amelioration in gastric secretion indices (pH and volume) and the marked decrease in the ulcer index. Moreover, SOIE could counteract the gastric oxidative stress induced by ethanol via a marked decline in malondialdehyde content as well as a significant (P < 0.05) increment in glutathione content and antioxidant enzymes activities (catalase and glutathione-s-transferase). Additionally, SOIE treatment caused a significant (P < 0.05) reduction in gastric nitric oxide content. Respecting morphological and histopathological studies, SOIE treatment at 200 mg/kg body weight caused marked healing of gastric lesions as indicated by no hemorrhagic bands or injuries observed as well as significantly reduced severity score of ulcer.ConclusionsSOIE could be used as a promising alternative antiulcerogenic compound to treat severe gastric lesions.

Ameliorative efficacy of Sepia officinalis ink extract on hepatorenal injury-induced following high-dose folic acid supplementation in rats

Sepia ink, a black suspension of melanin granules, is a multifunctional marine bioactive material. The present study aims to evaluate the ameliorative effect of the ink extract (IE) of the cuttlefish (Sepia officinalis) during high dosage administration of the FA in rats. Kidney injury induced by a single oral dose of FA (250 mg /kg). Eighteen male Wistar albino rats were the control, FA group, and FA+ IE group (250mg/kg). The IE showed a significant ameliorative effect against hepatorenal injury induced by high intake of FA as evident by decreasing the levels of serum aminotransferases (AST and ALT), urea, creatinine, uric acid, and significantly increased total serum albumin. Treatment with IE normalized the antioxidant status of the injured animals by reducing the MDA and the significant increase in the levels of GSH and CAT. The present study revealed that IE had an insightful effect against hepatorenal injury-induced following high intake of FA in rats, as it alleviates the alterations in the oxidative stress markers.

Ameliorative efficacy of Sepia officinalis ink extract on hepatorenal injury-induced following high-dose folic acid supplementation in rats

Ameliorative efficacy of Sepia officinalis ink extract on hepatorenal injury-induced following high-dose folic acid supplementation in rats

Ameliorative effect of ink extract and polysaccharide of Sepia officinalis on hepatotoxicity, renal toxicity and hematological disorders in adult male albino rats treated with cyclophosphamide

This study aimed to investigate ameliorative effects of the crude extract (SIE) and polysaccharide (SIP) of the ink of Sepia officinalis, on some biochemical and hematological disorders induced by cyclophosphamide (CP). Forty adult male albino Wistar rats were divided into five groups (n= 8 each). In the control group, rats were administered orally with 0.9% isotonic saline solution at a dose (5 ml/kg b.w.). All the other groups were i.p. injected with a single dose of CP (200mg/kg b.w.) only for one time. Then the third group was treated with oral administration of (SIE) (200mg/kg b.w.) daily for 60 days, the fourth group was treated with oral administration of (SIP) (80mg/kg b.w.) daily for 60 days and the fifth group was treated with oral administration of (SIE, 200mg/kg b.w. + SIP, 80mg/kg b.w.) daily for 60 days. All the animals were slaughtered by the end of the experiment for collecting the blood samples for hematological and biochemical assays. The biochemical results indicated that administration of CP was associated with a significant increase in the levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), urea, creatinine and uric acid. Moreover, a significant decrease in the levels of albumin and total protein was recorded. In addition, hematological disorders including a significant suppression on the numbers of RBCs, WBCs and PLTs, with a remarkable reduction in hemoglobin contents (Hb) and a significant drop in PCV values. Concomitant administration of SIE and SIP alleviated the altered biochemical and hematological parameters.

Quantitation of eumelanin and pheomelanin markers in diverse biological samples by HPLC-UV-MS following solid-phase extraction

Eumelanin and pheomelanin are well known and common pigments found in nature. However, their complex polymer structure and high thermostability complicate their direct chemical identification. A widely used analytical method is indirect determination using HPLC with UV detection of both types of melanin by their most abundant oxidation products: pyrrole-2,3-dicarboxylic acid (PDCA), pyrrole-2,3,5-tricarboxylic acid (PTCA), thiazole-4,5-dicarboxylic acid (TDCA), and thiazole-2,4,5-tricarboxylic acid (TTCA). An increasing interest in pigmentation in biological research led us to develop a highly sensitive and selective method to identify and quantify these melanin markers in diverse biological samples with complex matrices. By introducing solid-phase extraction (SPE, reversed-phase) following alkaline oxidation we could significantly decrease background signals while maintaining recoveries greater than 70%. Our HPLC-UV-MS method allows for confident peak identification via exact mass information in corresponding UV signals used for quantitation. In addition to synthetic melanin and Sepia officinalis ink as reference compounds eumelanin markers were detected in brown human hair and a brown bivalve shell (Mytilus edulis). Brown feathers from the common chicken (Gallus g. domesticus) yielded all four eumelanin and pheomelanin markers. The present method can be easily adapted for a wide range of future studies on biological samples with unknown melanin content.

The unique preservation of Sepia soft tissues in the Miocene deposits (Serravalian, Vienna Basin): Implications for the origin of microbodies in the fossil record

For the first time, we report an ancient cuttlefish ink sac possessing microbodies from the lower Serravallian (Middle Miocene) deposits of the Vienna Basin (Central Paratethys). We compare the geochemistry of these microbodies with the ink sac of the extant genus Sepia. The unique preservation of the cuttlefish soft tissue resulted from a high sedimentation rate and a low degree of sediment reworking and bioirrigation, which led to the dominance of anaerobic degradation pathways such as sulfate reduction below a thin oxic surface layer. The Recent Sepia melanin extracted from the ink sac is commonly used as a standard in biochemical analyses. However, it was unclear whether melanosomes can be preserved in fossil cuttlefishes. Although some portion of melanin-bearing microbodies is affected by pyritization, we show that some microbodies are still composed of eumelanin using Raman spectroscopy. They consist of particles corresponding to the size and shape of melanosomes observed in extant Sepia according to images produced with a scanning electron microscope. The Raman microspectroscopy of these microbodies in the Miocene Sepia shows identical chemical composition as in the Sepia officinalis ink melanin standard. Therefore, we confirm that these microbodies represent true fossil melanin pigment and melanosome remnants rather than being produced by microbes.

Eumelanin-releasing spongy-like hydrogels for skin re-epithelialization purposes

Melanin function in the skin has been associated with pigmentation but other properties such as electrical conductance, photoprotection, and antioxidant and antimicrobial activity have also been recognized. Nonetheless, the use of melanin in a skin wound healing context has never been considered. In this sense, eumelanin particles with a typical round and nano-sized morphology and electrical conductivity of 2.09 × 10−8 S cm−1 were extracted from the ink of Sepia officinalis. The ability of primary human keratinocytes (hKCs) to phagocyte eumelanin, which was then accumulated in cytosolic vesicles and nuclei surroundings, was demonstrated. Keratinocyte viability and maturation was not affected by eumelanin contact, but at eumelanin amounts higher than 0.1 mg l−1 cell morphology was altered and cell proliferation was inhibited. A time and eumelanin amount-dependent reduction of reactive oxygen species (ROS) released by eumelanin-containing ultraviolet (UV)-irradiated keratinocytes was observed. Eumelanin-containing gellan gum (GG) spongy-like hydrogels allowed a sustained release of eumelanin in the range of 0.1 to 5 mg l−1, which was shown in vitro to not be harmful to hKCs, and the absence of a strong host reaction after subcutaneous implantation in mice. Herein, we propose spongy-like hydrogels as sustained release matrices of S. officinalis eumelanin for predicting a beneficial role in skin wound healing through a direct effect over keratinocytes.

Sepia ink as a surrogate for colloid transport tests in porous media

We examined the suitability of the ink of Sepia officinalis as a surrogate for transport studies of microorganisms and microparticles in porous media. Sepia ink is an organic pigment consisted on a suspension of eumelanin, and that has several advantages for its use as a promising material for introducing the frugal-innovation in the fields of public health and environmental research: very low cost, non-toxic, spherical shape, moderate polydispersivity, size near large viruses, non-anomalous electrokinetic behavior, low retention in the soil, and high stability.Electrokinetic determinations and transport experiments in quartz sand columns and soil columns were done with purified suspensions of sepia ink. Influence of ionic strength on the electrophoretic mobility of ink particles showed the typical behavior of polystyrene latex spheres. Breakthrough curve (BTC) and retention profile (RP) in quartz sand columns showed a depth dependent and blocking adsorption model with an increase in adsorption rates with the ionic strength. Partially saturated transport through undisturbed soil showed less retention than in quartz sand, and matrix exclusion was also observed. Quantification of ink in leachate fractions by light absorbance is direct, but quantification in the soil profile with moderate to high organic matter content was rather cumbersome.We concluded that sepia ink is a suitable cheap surrogate for exploring transport of pathogenic viruses, bacteria and particulate contaminants in groundwater, and could be used for developing frugal-innovation related with the assessment of soil and aquifer filtration function, and monitoring of water filtration systems in low-income regions.

Therapeutic effect of Sepia ink extract against invasive pulmonary aspergillosis in mice

Invasive pulmonary aspergillosis (IPA) is a life-threatening disease in immunocompromised patients that requires aggressive therapy. Because of the widespread use of antibiotics, corticosteroids, antitumor drugs, and immunosuppressive drugs, the morbidity of IPA is currently increasing. The ink secretion of molluscan species was identified as one of the novel sources of bioactive compounds. So the present study designed to investigate the antifungal and antioxidant effects of Sepia officinalis ink extract against IPA in mice. Eighty neutropenic infected mice were randomly assigned into four main groups (20 mice/group). The 1st group was treated with saline, neutropenic infected, the 2nd group was treated with ink (200mg/kg) and the 3rd group was treated with amphotericin B (150mg/kg) and the 4th group was treated with ink plus amphotericin B (Ink 200mg/kg and AMB 150mg/kg). Treatment was started at 24h after fungal inoculation and was administered for 3 consecutive days. The present study demonstrated good in vitro and in vivo antifungal activity of IE against Aspergillus fumigatus. Compared with IPA group; IE-treated, AMB-treated, and AMB+IE-treated animals had a 67.80%, 83.41%, and 72.68% reduction in the pulmonary fungal burden, respectively. Treatment with IE and/or AMB for one and three days significantly decreased MDA and increased GSH and SOD levels in the lung tissues as compared with the infected untreated group. In conclusion, the results of our in vivo and in vitro studies demonstrate that IE has therapeutic effect against invasive pulmonary aspergillosis via reducing oxidative stress.

Toxicity of melanin-free ink of Sepia officinalis to transformed cell lines: identification of the active factor as tyrosinase

The melanin-free ink of the cephalopod Sepia officinalis is shown to contain a heat labile proteinaceous component toxic to a variety of cell lines, including PC12 cells. Gel filtration chromatography indicated that the toxic component was concentrated in those fractions eluted at a molecular weight higher than 100 kDa and exhibiting the highest tyrosinase activity. SDS–PAGE analysis of the active fractions displayed a single major band migrating at an approximate molecular weight of 100 kDa, identical with that of the single tyrosinase band in the melanin-free ink. These data unambiguously demonstrated the identity of the toxic component with tyrosinase. Treatment of purified Sepia as well as of mushroom tyrosinase with an immobilized version of proteinase K resulted in a parallel loss of tyrosinase activity and cytotoxicity. Sepia apotyrosinase was ineffective in inducing cytotoxicity in PC12 cells. Purified Sepia tyrosinase was found to induce a significant increase in caspase 3 activity in PC12 cells, leading eventually to an irreversible apoptotic process. Overall, these results disclose a hitherto unrecognized property of tyrosinase that may lead to a reappraisal of its biological significance beyond that of a mere pigment producing enzyme.

Molecular cloning of a peroxidase mRNA specifically expressed in the ink gland of Sepia officinalis

A full-length cDNA encoding a peroxidase has been assembled from two different overlapping cDNA clones isolated from the cDNA library of Sepia officinalis ink gland and 5′ RACE procedure. The cDNA of 2898 bp contains an open reading frame of 2778 bp corresponding to a protein of 926 amino acids. The protein shows 76% identity with a peroxidase isolated from the light organ of the squid Euprymna scolopes and approximately 30% identity with mammalian peroxidases. Northern blot analysis of mRNA from various tissues of Sepia officinalis reveals that peroxidase is exclusively expressed in the ink gland. These results represent the first characterization of a melanogenic peroxidase whose possible involvement in the biosynthesis of melanin is discussed.