Polyphenol-rich extracts of Terminalia laxiflora and Terminalia brownii give promising anti-Candida activity.

POLYPHENOL- RICH LEAF EXTRACTS OF TERMINALIA LAXIFLORA AND TERMINALIA BROWNII GIVE PROMISING ANTI-CANDIDAL ACTIVITY

Enass Yousif A Saliha,b,*, Markku Kanninenb, Riitta Julkunen-Tiittoc, Marketta Sipib, Olavi Luukkanenb, Heikki Vuorelaa, Pia Fyhrquista
aFaculty of Pharmacy, Division of Pharmaceutical Biosciences, Viikki Biocenter, PO Box 56, FIN-00014 University of Helsinki, Finland
MAIN CONCLUSION
The present study justifies the uses of Terminalia laxiflora and brownii leaves, roots and stem bark as water extracts and decoctions for the treatment of fungal infections in African traditional medicine. The antifungal leaf extracts of these species were found to be rich in ellagitannins and gallotannins as well some flavonoids which have not been reported before in these species.

INTRODUCTION
Terminalia laxiflora and Terminalia brownii occur in the tropical and subtropical savannah forest regions in Africa. Both species have a broad spectrum of medicinal uses in African traditional medicine [1]. In Sudan the water extracts and decoctions are used against malaria, cough as well as for bacterial and fungal infections [2, 3]. The fumigation of extracts of the stem wood and root are used customarily for treatment of skin and vaginal infections and for haemorrhoids [3]. Due to its homeostatic, stringent and healing effects, the fumigation is used to accelerate virginal operation recovery after delivery.

MATERIALS AND METHODS
Extracts of various polarities, obtained using sequential extraction and liquid partition were tested for their anti-Candida activity using agar diffusion and broth microdilution methods [4]. Human pathogenic Candida albicans, C. glabrata, C. tropicalis and C. parapsilosis were used for the screenings. Ethyl acetate extracts of the leaves of T. laxiflora and T. brownii were phytochemically analyzed using HPLC-DAD and UHPLC/QTOF-MS [5].

RESULTS AND DISCUSSION
All the tested polar leaf extracts of T. laxiflora gave good antifungal effects against C. glabrata, and especially large inhibition zones were obtained for an ethyl acetate extract, followed by a decoction, a cold water extract and a methanol extract (Figure 1). Ethyl acetate extracts of the root and stem bark of T. laxiflora gave promising growth inhibitory results against C. parapsilosis. The extracts were found to give less antifungal activity against C. albicans compared to ethyl acetate extract her species of yeast. The best activity against C. albicans was recorded for the methanolic Soxhlet extracts and EtOAc extracts of the root and hot water extracts of the stem wood of T. laxiflora. Ethyl acetate and methanolic Soxhlet extracts of the leaves of T. brownii gave inhibition zones of 23 and 21 mm against C. glabrata and C. tropicalis, respectively. Good antifungal results against C. glabrata were also obtained with an ethyl acetate extract and a hot water decoction of the stem bark of T. brownii.
We investigated the polyphenol composition of ethyl acetate extracts of the leaves of T. brownii and T. laxiflora since leaf extracts of these species have not been studied earlier in this context. Moreover, ellagitannins and gallotannins seem to dominate quantitatively and qualitatively in these extracts and their good antifungal activities are suggested partly to be due to these compounds. A high diversity of gallotannins could be seen in the leaf extracts of both species, T. laxiflora containing twenty-nine and T. brownii twenty-eight gallotannins. Pentagalloglucose at m/z 939.6700 (M-H)- and trigalloylglucose at m/z 635.0889 (M-H)- were present in both species. Ellagitannins were also present in a high variety and concentration in both species. Corilagin as well as five unknown ellagitannins, including an ellagitannin at m/z 617.0163 (M-H)- were identified in the leaves of T. laxiflora. Terflavin B, corilagin and ten unknown ellagitannins, among them an ellagitannin at m/z 541.0291 (M-H)-, were found in the leaves of T. brownii. The flavonoids quercetin 3-rhamnoside in T. laxiflora and apigenin, myricetin 3-rhamnoside and luteolin 7-glucoside in T. brownii have not been described earlier in these species. The high diversity of ellagitannins and gallotannins in the active extracts warrant further studies on their molecular structures.