EVALUATION OF HPLC, PHYTOCHEMICAL, ANTICHOLINESTERASE AND ANTIOXIDANT PROFILES OF THE AERIAL PARTS OF ASPERULA TAURINA SUBSP. CAUCASICA

Öz

Objective: In this study, we aimed to evaluate the HPLC, phytochemical, anticholinesterase and antioxidant profiles of the aerial parts of Asperula taurina subsp. caucasica.

Material and Method: The fingerprint of the phenolic compounds of the methanolic extract of the plant was obtained using RP-HPLC method. The method was also validated in terms of detection limits, quantification limits, linearity, accuracy, precision and selectivity. The phenolic contents of A. taurina subsp. caucasica were detected as proto-catechuic acid, p-OH benzoic acid and benzoic acid. In the phytochemical studies, quercetin 3-O-β-galactoside was isolated from the ethyl acetate subfraction of A. taurina subsp. caucasica using by several chromatographic methods. The structure of the pure compound was elucidated by means of spectral analysis (1H NMR, 13C NMR, and ESI-MS). Anticholinesterase and antioxidant activity studies were performed on quercetin 3-O-β-galactoside and the methanolic extract of the plant.

Result and Discussion: While quercetin 3-O-β-galactoside shown moderate inhibitory activity against butyrylcholinesterase at 200 μg/ml, quercetin3-O-β-galactoside and the metanolic exract of the plant did not show acetylcholinesterase inhibitory activity. Quercetin 3-O-β-galactoside shown DPPH free radical scavenging activity at 50 and 100 μg/ml, moderate lipid peroxidation inhibitory activity at 25, 50 and 100 μg/ml; the methanolic extract of the plant moderate lipid peroxidation inhibitory activity at 25, 50 and 100 μg/ml. In conclusion, A. taurina subsp. caucasica and quercetin 3-O-β-galactoside could be important and valuable sources for protecting our body health, especially Alzheimer’s disease.

Anahtar Kelimeler

• anticholinesterase activity
• antioxidant activity
• Asperula taurina subsp. caucasica
• fingerprint
• HPLC
• quercetin 3-O-β-galactoside
• Rubiaceae

ASPERULA TAURINA SUBSP. CAUCASICA'NIN TOPRAK ÜSTÜ KISIMLARININ YBSK, FİTOKİMYASAL, ANTİKOLİNESTERAZ VE ANTİOKSİDAN PROFİLLERİNİN DEĞERLENDİRİLMESİ

Öz

Amaç: Bu çalışmada, Asperula taurina subsp. caucasica'nın toprak üstü kısımlarının YBSK, fitokimyasal, antikolinesteraz ve antioksidan profillerinin değerlendirilmesi amaçlanmıştır.

Gereç ve Yöntem: Bitkinin metanolik ekstresinin fenolik bileşiklerinin parmak izi kromatogramı, geliştirilen RP-YBSK yöntemi kullanılarak elde edilmiştir. YBSK yöntemi saptama limitleri, nicelik sınırları, doğrusallık, doğruluk, hassaslık ve seçicilik açısından valide edilmiştir. A. taurina subsp. caucasica fenolik içeriği, protokatekuik asit, p-OH benzoik asit ve benzoik asit olarak tespit edilmiştir. Fitokimyasal çalışmalarda ise, çeşitli kromatografik yöntemler kullanılarak A. taurina subsp. caucasica etil asetat alt fraksiyonundan kersetin 3-O-β-galaktozit izole edilmiştir. Saf bileşiğin yapısı, spektrum analizi (1H NMR, 13C NMR ve ESI-MS) yardımıyla aydınlatılmıştır. Kersetin 3-O-β-galaktozit ve bitkinin metanolik ekstresi üzerinde antikolinesteraz ve antioksidan aktivite çalışmaları yapılmıştır.

Sonuç ve Tartışma: Kersetin 3-O-β-galaktozit, butirilkolinesteraz'a karşı 200 μg/ml'de orta düzeyde inhibe edici aktivite gösterirken, kersetin 3-O-β-galaktozit ve bitkinin metanolik ekstresi asetilkolinesteraz inhibitör etki göstermemiştir. Kersetin 3-O-β-galaktozit, 50 ve 100 μg/ml'de DPPH serbest radikal süpürücü aktivite, 25, 50 ve 100 μg/ml'de orta derecede lipit peroksidasyon inhibisyon aktivitesi gösterirken; bitkinin metanolik ekstresi 25, 50 ve 100 μg/ml'de orta düzeyde lipit peroksidasyonu önleyici aktivite göstermiştir. Sonuç olarak, A. taurina subsp. caucasica ve kersetin 3-O-β-galaktozit, özellikle Alzheimer hastalığından korunmada önemli ve değerli bir doğal ürün kaynağı olabilir.

Anahtar Kelimeler

• antikolinesteraz aktivite
• antioksidan aktivite
• Asperula taurina subsp. caucasica
• kersetin 3-O-β-galaktozit
• parmak izi
• Rubiaceae
• YBSK

Kaynakça

1. Evans, W.C. (1989). Trease and Evans’ Pharmacognosy (13th Ed), BailliereTindall, London.
2. Burnett, A.R., Thomson, R.H. (1968). Naturally occurring quinones Part XII. Antraquinones and related naphtalenic compounds in Galium ssp. and in Asperula odorata. Journal of the Chemical Society, 0, 854–857.
3. Itokawa, H., Ibraheim, Z.Z., Qiao, Y.F, Takeya, K. (1993). Antraquinones, naphthohydroquinones and naphtahydroquione dimers from Rubia cordifolia and their cytotoxic activity. Chemical and Pharmaceutical Bulletin, 41, 1869–1872.
4. Adesogan, E.K. (1973). Anthraquinones and anthraquinols from Morinda lucida. Tetrahedron, 29, 4099–4102.
5. Inouye, H., Takeda, Y., Nishimura, Y.H., Kanomi, A., Okuda, T., Puff C. (1988). Chemotaxonomic studies of Rubiaceous plants containing iridoid glycosides. Phytochemistry, 27, 2591–2598.
6. Borisov, M.I. (1974). Coumarins of the genus Asperula and Galium. Khimiya Prirodnykh Soedinenii, 10, 82.
7. Itokawa, H., Qiao, Y.F., Takeya, K., Iitaka, Y. (1989). New triterpenoids from Rubia cordifolia var. pratensis (Rubiaceae). Chemical and Pharmaceutical Bulletin, 37, 1670–1672.
8. Borisov, M.I., Serbin, A.G. (1972). Flavonoids of Asperula oppositifolia. Khimiya Prirodnykh Soedinenii, 8, 122–123.

9. Ozturk, M. (2013). Asperula anatolica (Rubiaceae), a new species from southeast Anatolia, Turkey. Turkish Journal of Botany, 37, 4654.
10. Ehrendorfer, F, Schöbeck-Temesy, E. (1982). Asperula L. in Flora of Turkey and the East Aegean Islands (Vol. 7), Davis, P.H. (Ed.), University Press, Edinburgh, 734–767.
11. Baytop, T. (1984). Therapy with Medicinal Plants in Turkey (Past and Present). Istanbul University Publications, Istanbul, 417.
12. Ozgen, U., Kazaz, C., Seçen, H., Coskun, M. (2006). Phytochemical Studies on the underground parts of Asperula taurina subsp. caucasica. Turkish Journal of Chemistry, 30, 15–20.
13. Kırmızıbekmez, H., Tiftik, K., Kúsz, N., Orban-Gyapai, O., Zomborszki, Z., Hohmann, J. (2017). Three New Iridoid Glycosides from the Aerial Parts of Asperula involucrate. Chemistry and Biodiversity, 14, 1–7.
14. Kırmızıbekmez, H., Bardakcı, H., Masullo, M., Kamburoglu, Ö., Erylmaz, G., Akaydn, G., Yesilada, E., Piacente, S. (2014). Flavonol glycosides and iridoids from Asperula lilaciflora. Helvetica Chimica Acta, 97, 1571–1576.
15. Sergeevna, I.N., Ilyina Tatyana, V., Mihaylovna, K.A., Leonidivna, T.E., Aleksandrovna, K.I. (2015). The Antihypoxic and Sedative Activity of the Dry extract from Asperula odorata L. Pharmacognosy Communications, 5, 233–236.
16. Kayalar, H., Kalyoncu, F., Minareci, E., Kalyoncu, F., Ergonul, B., Kayalar, H. (2011). Chemical Compositions and Antioxidant Activities of Five Endemic Asperula Taxa. Archives of Biological Sciences, 63, 537–543.
17. Kim, D.O., Lee, C.Y. (2004). Comprehensive study on vitamin C equivalent antioxidant capacity (VCEAC) of various polyphenolics in scavenging a free radical and its structural relationship. Critical Reviews in Food Science and Nutrition, 44, 253–273.
18. Ellman, G.L., Courtney, K.D., Andres, V., Featherston, R.M. (1961). A new and rapid colorimetric determination of acetylcholinesterase activity. Biochemical Pharmacology, 7, 88–95.
19. Blois, M.S. (1958). Antioxidant determinations by the use of a stable free radical. Nature, 181, 1199-1200.
20. Peres, R.G., Tonin, F.G., Tavares, M.F., Rodriguez-Amaya, D.B. (2013). HPLC-DAD-ESI/MS identification and quantification of phenolic compounds in Ilex paraguariensis beverages and on-line evaluation of individual antioxidant activity. Molecules, 18, 3859–3871.
21. Miller, H.E. (1971). A simplified method for the evaluation of antioxidants. Journal of the American Oil Chemists' Society, 48, 91.
22. Otsuka, H., Yoshimura, K., Yamasaki, K., Cantoria, M.C. (1991). Isolation of 10-acyl iridoid glucosides from a Philippine medicinal plant, Oldenlandia corymbosa L. (Rubiaceae). Chemical and Pharmaceutical Bulletin, 39, 2049–2052.
23. International Conference on Harmonization (ICH) (2005). Validation of Analytical procedurs-PA/PH/OMCL (05) 47 DEF, elaborated by OMCL Network/EDQM of the Council of Europe.