Introduction

Ocean is the rich biodiverse and it has occupied 70% of the earth. The marine environment needs more adaptive changes in the organisms so that they form a special structure of the bioactive substances. Marine organisms are living in complex habitats and are exposed to extreme conditions, such as temperature, salinity, pressure, and illumination; thus, they could produce a wide variety of secondary metabolites (Hamed et al. 2015). Ascidians belongs to the phylum of Chordata, which encompasses all vertebrate animals, including mammals and, therefore, they represent the most highly evolved group of animals commonly investigated by marine natural products chemists (Menna et al. 2011). Recently marine peptides have opened a new perspective for novel and efficient drug developments. Cyclic and linear peptides are derived from marine ascidians have been increased our knowledge regarding novel cytotoxic, antimicrobial, anti-oxidant, antiviral and many other biological properties (Destoumieux et al. 1997; Li et al. 2014; Cheung et al. 2015). It has the advantages of less toxicity, high targeting specificity and effective biological activity. These bioactive peptides are extracted from sponges, ascidians, seaweeds, and mollusks and reported to have various pharmacological properties. The bioactive peptides often have 3 to 20 amino acid residues, and their biological activities are determined based on their sequence strength and amino acid composition (Pihlanto-Leppälä 2000; Bhat et al. 2015; Stoye et al. 2016). Recently, these peptides have been paid attention to unravel their structure, sequences and compositional properties (Lordan et al. 2011). The objectives of this review are to provide an overview of the available sources of marine bioactive peptides and their biological activities and its potential applications in functional foods and pharmaceuticals for further drug development.

Techniques for the Synthesis of Bioactive Peptides

The widely used protocols for the isolation of bioactive peptides are by chemical methods containing acid and alkaline hydrolysis, enzymatic hydrolysis is performing on the laboratory scale and Recombinant DNA technology (Guzmán et al. 2006; Magana et al. 2015; Lemes et al. 2016). The few other common methods were used by bioreactor systems or immobilized enzymes to increase product yield and fermentation technology. Recently, many advanced extraction techniques like supercritical fluid extraction, pressurized solvent extraction, microwave-assisted extraction, ultrasound-assisted extraction, pulsed electric field-assisted extraction and enzyme-assisted extraction are developed and most preferred for extracting bioactive peptides (Grosso et al. 2015). The acid hydrolysis is an important method for chemical modification that can significantly change the structure and functional properties of peptides and this method is preferred over other pretreatments because of its low cost and higher effectiveness (Lee and Jeffries 2011; Loow et al. 2016). The enzymatic methods of peptide bond formation are mediated by an enzyme (protease) in free or immobilized form and this method is especially useful in the synthesis of very short peptides (2–5 oligomers) as well in the condensation of large peptide fragments (So et al. 1998; Perez Espitia et al. 2012). The recombinant DNA technology offers modern methods of cloning and gene expression in microorganism and other organisms, allowing production of a several recombinant peptides (Sewald and Jakubke 2002). While comparing the all available methods for the synthesis of bioactive peptides, the recombinant DNA technology approach seems to be the most cost effective and very good alternative for large-scale peptide production (Li 2011).

Purification and Characterization of Peptides

The separation and purification of bioactive peptides are primarily based on their physical and chemical characteristics, such as their molecular size, charge, polarity, solubility, and specific covalent or noncovalent interactions (Jo et al. 2017). The separation strategies are mainly employed depending on the characteristics feature the target peptides. The different techniques used in the purification of peptides are ion-exchange chromatography, affinity chromatography, capillary electrophoresis, size exclusion chromatography and reverse-phase high-performance liquid chromatography (RP-HPLC). The isolated and purified peptides are characterized with different analytical equipment, majorly as LC-MS, LC/-MS/MS or LC-NMR (Martins et al. 2014; Pérez-Victoria et al. 2016; Ulrike 2016) and fast atom bombardment mass spectrometry (FAB-MS), electrospray ionization (ESI-MS), or matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) (Jo et al. 2017). The accuracy of these new techniques helped to determine the molecular mass to charge ratio of the peptide, along with its chemical structure, with high resolution and sensitivity. In future, genome mining will be one of the most important aspects and will provide useful information. Generally, the silent genes/gene clusters, is the key factor to discover the interesting and new peptide compounds (Gross 2009).

Physiological Properties of Bioactive Peptides Derived from Ascidians

Bioactive peptides from ascidians has three types of structure including cyclic, acyclic (depsipeptides), and linear that have potential biological functions of antineoplastic, antiviral activity, antidiabetic, antioxidant activity, immunomodulatory properties (Schwartsmann et al. 2001). The Urochordata, also known as tunicates and ascidians, have emerged as a rich source of peptides and successfully preceded several drugs candidates including few under the clinical trials (Table 1). Lee et al. (1997 ) discussed that antimicrobial peptides of Clavanins A, B, C and D contain 23 amino acid residues were reported from Solitary tunicate, Styela clava and it has shown antibacterial activity against the three Gram-negative (E. coli, S. typhimurium, P. aeruginosa) and two Gram-positive strains (S. aureus, M. flavus) at 0.4–5 μM concentration (MIC).

Table 1 Successful ascidian marine peptides in clinical development
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Halocyamine A and B are two antimicrobial tetrapeptides isolated from the hemocytes of the ascidian Halocynthia roretzi (Table 2). It shows antimicrobial activities against several strains of bacteria and yeasts with cytotoxic activities of neuroblastoma N-18 cells at 160 µM and human hepatoma Hep-G2 cells at 100 µM (Azumi et al. 1990). The cytotoxic depsipeptides of Tamandarins A and B have been discovered from a marine ascidian of the family Didemnidae, which was evaluated against different human cancer cell lines such as pancreatic carcinoma (BX-PC3), prostate carcinoma (DU145) and the head and neck carcinoma (UMSCC10b). Tamandarin A showed a 50% reduction in overall cell survival (IC50) at a concentration of 1.79, 1.36, and 0.99 ng/mL, respectively (Hamad and Shioiri 2005; Vervoort et al. 2000). Styelin D, the antimicrobial peptides with a 32-residue C-terminus, extracted from ascidian Styela clava blood cells (hemocytes), contains two novel amino acids, dihydroxyarginine and dihydroxylysine (Vervoort et al. 2000). Lee et al. 2001 reported antimicrobial peptides were isolated from the solitary tunicate Halocynthia aurantium compound of Dicynthaurin is composed two 30-residue cynthaurin monomers containing only four lysines and no arginines or histidines and its show activity against the 5 bacterial pathogens (E. coli, P. aeruginosa, M. luteus, L. monocytogenes, S. aureus) at 140 μg/mL (MIC). Perez and Faulkner (2003) reported that bistratamides C–J, cyclic hexapeptides derived from Philippines ascidian Lissoclinum bistratum showed moderate cytotoxicity against HCT-116 cell line at varying concentrations of 7.9; 28; 5; 1.7; 9;1 μg/mL. Styelin D is effective against Gram-positive and Gram-negative bacteria, even in excessive salinity (200 mM NaCl). These antibiotic properties are used for the development of microbicides effective at high concentrations of NaCl or physiological conditions (Aneiros and Garateix 2004).

Table 2 Peptides derived from marine ascidians with possible therapeutic applications
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A new cytotoxic cyclic heptapeptide, Mollamide (Fig. 1c) obtained from the ascidian Didemnum molle, shows very good cytotoxicity effect against various cell lines with IC50 values of 1 μg/mL towards P388 murine leukemia line, 2.5 μg/mL against A549 human lung carcinoma and HT29 human colon carcinoma CV1 (monkey kidney fibroblast) cell lines with IC50 value of 2.5 μg/mL (Carroll et al. 1994). Donia et al. 2008 reported that two new mollamides B and C isolated from the Indonesian tunicate Didemnum molle. Mollamides B evaluated against different cell lines of non-small cell lung cancer cell line (H460), breast cancer cell line (MCF7) and CNS cancer cell line (SF-268) cytotoxicity activity at 100 μM. The antimalarial activity was assessed against Plasmodium falciparum (D6 clone and W2 clone) at IC50 values shows of 2.0 and 2.1 μg/mL. From Mollamides B showed marginal activity against Leishmania donoVani at IC50 and IC90 values of 18 and 35 μg/mL. The cytotoxicity against HIV-1 in human PBM cells at EC50 value of 48.7 μM in vitro condition. Mollamide C showed minor activity against the two leukemias, five solid tumors, murine, human normal cell and other few cell lines. Trunkamide A is one of cyclopeptide with a tiazoline ring similar to Mollamide isolated from ascidians of the genus Lissoclinum, where antitumor activities under pre-clinical trials are being demonstrated (Hamada and Shioiri 2005; Wipf et al. 1995).

Fig. 1
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a–d Important peptides isolated from the ascidians

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Fu et al. 1998 isolated the four new cyclic peptides, patellamide G and ulithiacyclamides E–G, along with the known patellamides A–C and ulithiacyclamide B from the ascidian Lissoclinum patella collected in Pohnpei, Federated States of Micronesia and it has shown vinblastine against the drug resistant cells (CEM/VBL100) was IC50 at 90 nM and Patellamides B and C exhibited in vitro modulation of multidrug resistance in CEM/VBL100 cells. The bicyclic peptide, Vitilevuamide derived from two marine ascidians, Didemnum cuculiferum and Polysyncranton lithostrotum and showed cytotoxic activity in several human tumor cell lines with LC50 values of 6, 124, 311 and 311 nM were obtained for human colon tumor (HCT116), lung cancer (A549), malanoma tumor (SK Mel-5) and kidney cancer cell lines (A498), respectively. Vitilevuamide suppressed tubulin polymerization and it can arrest the cell cycle of target cells in the G2/M phase (Edler et al. 2002).

Two new antimicrobial peptides, Halocyntin and papillosin were derived from hemocytes of the solitary tunicate, Halocynthia papillosa and these molecules showed the antibacterial activity against Gram-positive and Gram-negative bacteria at 0.75–100 μM (MBC) (Galinier et al. 2009). Five new lipopeptides, peptidolipins B–F were derived from the associated organism of Nocardia sp. isolated from the ascidian Trididemnum orbiculatum and Peptidolipins B and E has shown the moderate antibacterial activity against MRSA and methicillin-sensitive Staphylococcus aureus (Wyche et al. 2012). Ko et al. 2016 discussed the antihypertensive Peptides of Ala-His-Ile-Ile-Ile were derived from the Tunicate Styela clava and showed Peptide increases glucose uptake via activation of the AMPK pathway and may potentially emerge as a novel target for the treatment of type II diabetes. Two cyclic Hexapeptides of Bistratamide M and N derived from the marine ascidian Lissoclinum bistratum were collected from Raja Ampat (Papua Bar, Indonesia) and it has shown the cytotoxic activity Human tumor cell lines of breast (MDA-MB-231), colon (HT-29), lung (A-549) and pancreas (PSN1) with LC50 values at ≥ 20 µm (Urda et al. 2017).

Recent Pharmacological Advances for Drug Development

The success of biotechnology in pharmaceutical industry has been developed, now by the extensive number of scientific advances for the currently approved novel products for human health care. In this concept various techniques are involved in the drug development. The confluence of computational and high-throughput experimental methods for protein-engineering and platform technologies has ushered in unprecedented opportunities to develop safe, effective, and more convenient protein and peptide therapeutics and also Recombinant DNA technology allow the production of a wide range of peptides, proteins and biochemicals from naturally nonproducing cells (Kumar et al. 2006; Torchilin 2008; Carter 2011). This technology, now approximately 25 years old, is becoming one of the most important technologies for currently drug development. Intelligent coupling of cellular tests with omics technologies will help to identify interesting drug candidates, to clarify mode of action and to avoid late stage failure of compounds in the pipeline (Fig. 2). More attention should be given to early investigation of toxicological risks and to pharmacokinetic behavior of the drug candidates. Modern biotechnology and pharmaceutical analysis has evolved beyond this to encompass combination techniques, high-throughput technologies, chemo metrics, micro dosing studies, miniaturization and nanotechnology. These recent techniques are very useful to ascidians peptides for future drug development.

Fig. 2
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Modern route for drug development from bioactive peptides: modern biotechnology

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Conclusion

In conclusion, bioactive peptides derived from marine ascidians have a potential for human health care. The unique chemical classes of peptides found in the ascidians have promising biological activities which make them excellent candidates for drug design and development. The ascidians peptides are important for pharmaceutical studies and discovering new therapeutic treatment like anti-tumor/anti-cancer, anti-HIV, antimicrobial, and various diseases. In this review, we concluded Marine ascidians contains a wide range of secondary metabolites such as protein, peptides and minor metabolites might serve as vital role in future drug development.