Mert Karış

Biological activities of skin and parotoid gland secretions of bufonid toads (Bufo bufo, Bufo verrucosissimus and Bufotes variabilis) from Turkey

Toad glandular secretions and skin extractions contain numerous natural agents which may provide unique resources for novel drug development. Especially the skin-parotoid gland secretions of toads from genus Bufo contain as many as 86 different types of active compounds, each with the potential of becoming a potent drug. In the present study, crude skin-parotoid gland secretions from Bufo bufo, Bufo verrucosissimus and Bufotes variabilis from Turkey were screened against various cancer cells together with normal cells using MTT assay. Furthermore, the antimicrobial properties of skin secretions were tested on selected bacterial and fungal species for assessing the possible medical applications. Antimicrobial activity of skin secretions was studied by determining minimal inhibitory concentration (MIC) in broth dilution method. Hemolytic activity of each skin-secretion was also estimated for evaluating pharmaceutical potential. Both skin-parotoid gland secretions showed high cytotoxic effect on all cancerous and non-cancerous cell lines with IC50 values varying between <0.1μg/ml and 6.02μg/ml. MIC results of antimicrobial activity tests were found to be between 3.9μg/ml and 250μg/ml. No hemolytic activities on rabbit red blood cells at concentrations between 0.5μg/ml and 50μg/ml were observed. In conclusion, skin-parotoid secretions of bufonid toads might be remarkable candidates for anti-cancer and antimicrobial agents without hemolytic activities.

Comparative Venomics of the Vipera ammodytes transcaucasiana and Vipera ammodytes montandoni from Turkey Provides Insights into Kinship

The Nose-horned Viper (Vipera ammodytes) is one of the most widespread and venomous snakes in Europe, which causes high frequent snakebite accidents. The first comprehensive venom characterization of the regional endemic Transcaucasian Nose-horned Viper (Vipera ammodytes transcaucasiana) and the Transdanubian Sand Viper (Vipera ammodytes montandoni) is reported employing a combination of intact mass profiling and bottom-up proteomics. The bottom-up analysis of both subspecies identified the major snake protein families of viper venoms. Furthermore, intact mass profiling revealed the presence of two tripeptidic metalloprotease inhibitors and their precursors. While previous reports applied multivariate analysis techniques to clarify the taxonomic status of the subspecies, an accurate classification of Vipera ammodytes transcaucasiana is still part of the ongoing research. The comparative analysis of the viper venoms on the proteome level reveals a close relationship between the Vipera ammodytes subspecies, which could be considered to clarify the classification of the Transcaucasian Nose-horned Viper. However, the slightly different ratio of some venom components could be indicating interspecific variations of the two studied subspecies or intraspecies alternations based on small sample size. Additionally, we performed a bioactivity screening with the crude venoms against several human cancerous and non-cancerous cell lines, which showed interesting results against a human breast adenocarcinoma epithelial cell line. Several fractions of Vipera a. transcaucasiana demonstrated a strong cytotoxic effect on triple negative MDA MB 231 breast cancer cells.

Geographic nestedness of Lyciasalamandra billae (Amphibia: Salamandridae) populations within L. antalyana and description of a new subspecies

A new subspecies of the Bille’s Lycian Salamander Lyciasalamandra billae is described from four localities in the vicinity of Geyikbayɪrɪ nearby Antalya, Turkey. It is distinguished from the nominotypical subspecies by colouration and surface pattern. The distribution area of the new subspecies is nested within the range of Lyciasalamandra antalyana, with L. a. gocmeni bordering in the north and the range of L. a. antalyana bordering in the south. New localities of both L. antalyana subspecies are reported. http://www.zoobank.org/urn:lsid:zoobank.org:pub:8F541E98-61F3-4EDD-A432614B0A250711

Molecular and morphological evidence for a new subspecies of Fazila’s Lycian Salamander Lyciasalamandra fazilae in South-west Anatolia

A new subspecies of the Fazila’s Lycian Salamander Lyciasalamandra fazilae is described based on material from ten localities in the Köyceğiz, Ortaca and Dalaman area in south-western Turkey. It is distinguished from the nominotypical subspecies by differences in the colouration pattern, morphometry and the mitochondrial molecular marker 16S rRNA. The distribution area of the new subspecies is located mainly in the western part of Dalaman River except for two seemingly intermediate populations (Şerefler and Sarsala-Kapıkargın). New localities for the species are reported.

Major biological activities and protein profiles of skin secretions of Lissotriton vulgaris and Triturus ivanbureschi

Abstract Objective The aim of this study was to determine the total protein amounts, protein profiles, in vitro cytotoxicities, antimicrobial activities and hemolytic effects of skin secretions of the Lissotriton vulgaris and Triturus ivanbureschi. Methods Skin secretions were obtained, clarified, supernatants snap-frozen then lyophilized. Total protein amounts were determined by BCA assay kit. Protein profiles were revealed by the SDS-PAGE. The cytotoxicity and antimicrobial activity were determined by using MTT assay and minimum inhibitory concentration (MIC) method. Hemolytic effects were measured on rabbit red blood cells. Results Lissotriton vulgaris and T. ivanbureschi skin secretions have totally 18 and 20 protein fractions. IC50 values were detected between 1.40 and 40.28 μg/mL. The MIC results were found between 7.8 and 250 μg/mL. Lissotriton vulgaris skin secretion showed low hemolytic effect while T. ivanbureschi skin secretion showed high hemolytic effect. Conclusion This study is the first report showing the potential of L. vulgaris and T. ivanbureschi skin secretions for cytotoxicity, antimicrobial and hemolytic activity as an alternative therapeutic approach for traditional uses. Further studies need to focus on purification of the active components from these skin secretions and mode of action on cancer cell lines and microorganisms as anti-agents.

Diyabetik sıçanların karaciğer ve böbrekleri üzerinde Caralluma tuberculata’nın hipoglisemik ve hipolipidemik etkisi ve güvenliği / Liquidambar orientalis Mill. reçine ekstraktlarının antioksidan, sitotoksik ve iNOS aktiviteleri

Abstract Objective: The aim of this study is to investigate the in vitro cytotoxicity and iNOS (inducible nitric oxide synthase) inhibitory, and antioxidant activity in order to assess the traditional usage of Liquidambar orientalis Mill resin extract. Methods: Different solvent extracts of Liquidambar orientalis Mill resin were prepared. The cytotoxicity of extracts was determined using MTT (3-(4,5-dimethyl-2-thiazolyl)- 2,5-diphenyl-2H-tetrazoliumbromide) assay. HeLa (Human cervix adenocarcinoma), A-549 (Human alveolar adenocarcinoma), MCF-7 (Human breast adenocarcinoma), CaCo-2 (Human colon colorectal adenocarcinoma), mPANC96 (Human pancreas adenocarcinoma), PC-3 (Human prostate adenocarcinoma), U87MG (Human glioblastoma- astrocytoma) and as a normal cell line HEK293 (Human embryonic kidney cells) and Vero (African green monkey kidney epithelial cells) were used for testing cytotoxicity. RAW 264.7 (murine macrophage cell lines) was used to determine the inhibition levels of inducible nitric oxide synthase (iNOS). HL-60 (human acute myeloid leukemia) was used to determine antioxidant activity as DCF production per cent. Results: Hexane, dichloromethane, methanol and water extracts were prepared, and their iNOS inhibitory, cytotoxic and antioxidant activity were investigated. The estimated IC50 values of extracts varied from 6.68 to 48.90 μg/ ml after treatment with different doses of extracts for 48 h. Inhibition of the hexane, dichloromethane, methanol, and water extracts on LPS-induced NO production in RAW 264.7 macrophage were showed that the all extracts inhibited NO production in activated RAW 264.7 cells, except methanol extract. Hexane, dichloromethane and methanol extracts inhibited NO production with IC50 values of 22 μg/ml, 22 μg/ml, and 21/ml μg, respectively. The antioxidant activity was determined by the production of DCF in HL-60 cells. The extracts showed a potent antioxidant effect, with an IC50 of in between from 19.01 to 39.77 μg/ml. Conclusion: This study is the first report showing the potential of Liquidambar orientalis Mill resin extracts for cytotoxicity, iNOS inhibition and the antioxidant activity as an alternative therapeutic approach for traditional uses. The results demonstrated that Liquidambar orientalis dichloromethane resin extracts showed strongest cytotoxic effect while all extracts except methanolic extracts exhibited moderate iNOS inhibition. All extracts other than hexane have a potent antioxidant effect in the cellular-based assay. In conclusion, further studies should focus on the purification of the active components of this extracts and to investigate the possible mode of action to obtain a better understanding of their potential use as cytotoxic, anti-inflammatory and antioxidant agents. Özet Amac: Bu calışmanın amacı, Liquidambar orientalis recine ekstresinin geleneksel kullanımını değerlendirmek uzere in vitro sitotoksisite, iNOS (induklenebilir nitrik oksit sentaz) inhibisyonu ve antioksidan aktivitelerinin araştırılmasıdır. Metod: Liquidambar orientalis recinesinin farklı cozgenlerde ekstreleri hazırlandı. Ekstrelerin sitotoksisitesi MTT (3-(4,5-dimetil-2-tiyazolil)-2,5-difenil-2H-tetrazolyumbromid) deneyi kullanılarak saptandı. HeLa (insan serviks adenokarsinomu), A-549 (insan alveolar adenokarsinomu), MCF-7 (insan goğus adenokarsinomu), CaCo-2 (insan kolorektal adenokarsinomu), mPANC96 (insan pankreas adenokarsinomu), PC-3 (insan prostat adenokarsinomu), U87MG (insan glioblastom-astrositom hucre hattı), normal hucre hattı olarak HEK293 (insan embriyonik bobrek hucreleri) ve Vero (Afrika yeşil maymun bobrek epitelyal hucreleri) sitotoksisite testi icin kullanıldı. RAW 264.7 (fare makrofaj hucre hattı), induklenebilir nitrik oksit sentaz inhibisyon seviyelerini belirlemek icin kullanıldı. HL-60 (insan akut miyeloid losemi) yuzde DCF uretimi olarak antioksidan aktivitesini belirlemek icin kullanıldı. Bulgular: Hekzan, diklorometan, metanol, su ekstreleri hazırlandı ve bunların iNOS inhibisyonu, sitotoksik ve antioksidan aktiviteleri araştırıldı. Sitotoksisite sonucu olarak, hucre hatlarının, ekstraktların farklı dozları ile muamelesinden 48 saat sonra hesaplanan IC50 değerleri 6.68-48.90 μg/ml arasında değişkenlik gostermektedir. iNOS inhibisyonu RAW 264.7 hucrelerinin lipopolisakkarit ile induklenmesi ile gercekleştirildi. Aktive olan macrofaj hucrelerindeki nitrik oksit (NO) uretiminin su ekstraktı dışındaki diğer ekstraktlar tarafından inhibe edildiği saptandı. Hekzan, diklorometan ve metanol ekstraktlarının NO uretim inhibisyonu IC50 değerleri sırasıyla, 22, 22, 21 μg/ml olarak bulunmuştur. Antioksidan aktivite, HL-60 hucrelerinde DCF uretimi ile tespit edilmiştir. Ekstraktlar, 19.01-39.77 μg/ml arasındaki IC50 değerleri ile potansiyel antioksidan etki gostermiştir. Sonuc: Bu calışma, Liquidambar orientalis recine ekstraktlarının geleneksel kullanımdaki alternatif terapotik potansiyellerini sitotoksisite, iNOS inhibisiyonu ve antioksidan aktivite acılarından ortaya koyan ilk rapor niteliğindedir. Eldeki veriler sonucunda en guclu sitotoksik aktiviteyi diklorometan ekstraktı gostermiştir. Metanolik ekstrakt dışındaki ekstraktlar orta duzeyde iNOS inhibisyonu ve hekzan ekstraktı dışındakiler potensiyel antioksidan etki gostermişlerdir. Sonuc olarak, gelecek calışmalarda ekstrakların potensiyel sitotoksik, antienflamatuvar ve antioksidan ajanlar olarak kullanımı icin aktif bileşenlerinin saflaştırılması ve olası etki mekanizmalarının belirlenmesine odaklanılmalıdır.