Hop and Medicinal Plants, Year XXVI, No. 1-2, 2018 ISSN 2360-0179 print, ISSN 2360-0187 electronic GALIUM SPECIES - POLYPHENOLIC CONTENT AND THEIR ANTIOXIDANT POTENTIAL HANGANU Daniela1, Ramona Flavia BURTESCU2, Sorina PETRESCU2, Flavia Roxana PRIPON FURTUNA2, Elisabeta CHIȘE3, Violeta TURCUȘ3,4, Daniela BENEDEC1*, Ilioara ONIGA1, Neli Kinga OLAH2,3 1„Iuliu Hațieganu” Medicine and Pharmacy University, Faculty of Pharmacy, Pharmacognosy Department, Cluj-Napoca, Romania 2SC PlantExtrakt SRL, Rădaia, jud. Cluj, Romania 3„Vasile Goldiș” Western University from Arad, Faculty of Pharmacy, Arad, Romania 4Romanian Academy, Institutul Naţional de Cercetări Economice „Costin C. Kiriţescu” National Institute for Economic Research, Mountain Economic Center CE-MONT Vatra Dornei, Romania *Corresponding authors: [email protected] Abstract. The genus Galium is well represented in the spontaneous Romanian flora. Three Galium species are more commonly used in therapy, Galium aparine L. or cleavers, Galium odoratum (L.) Scop. or woodruff and Galium verum L. or lady’s bedstraw. Tinctures preparated from aerial part of these species was chemically analyzed and their antioxidant activity was evaluated. The total phenylpropane derivatives and total flavonoids contents were determined by spectrophotometrically methods. The antioxidant capacity was evaluated in vitro by FRAP, CUPRAC and xanthine oxidase inhibition methods. The results showed that the tinctures were rich in polyphenols and had a strong antioxidant capacity, which is confirmed by all used methods. A good correlation between the antioxidant capacity and the content of polyphenols could be highlighted. Keywords: antioxidant capacity, Galium aparine L., G.odoratum (L.) Scop., G. verum L., polyphenols Introduction Genus Galium is part of the Rubiaceae family, comprising more than 600 species of grassy, annual or perennial plants, specific to the temperate zone, of both, northern and southern hemispheres. According to an evaluation the number of species of Galium is between 629 and 650, some of which can be easily confused with Sherardia arvensis L. or species of the 84 genus Asperula. Sometimes the species Galium odoratum L.-woodruff is even included in the genus Asperula under the name Asperula odorata L. (Ardelean and Mohan, 2008; Ciocârlan, 2009). In Romania, genus Galium or bedstraw, has a special meaning being represented by around 38 species, including Galium aparine L. called cleavers or stickyweed, Galium mollugo L. named also hedge bedstraw, Galium odoratum (L.) Scop., Galium purpureum DC. or reddish bedstraw, Galium ruboides L. or European bedstraw, Galium schultesii West. forest bedstraw or Galium verum L. named also lady’s bedstraw (Ardelean and Mohan, 2008; Ciocârlan, 2009). Galium aparine L. or the cleavers, named also stickyweed, is an annual grassy plant with stem and leaves covered by short hair. The plant is widespread in Europe, North Africa and Asia. The rectangular stem is wearing simple leaves, almost oblanceolate or linear, attached 6-8 in form of the whorls. The flowers are small, star-shaped, white or greenish, grouped by 2-3. The globular, thorny fruits contain 1 to 3 reunited seeds. It is considered an invasive plant in certain regions, with an irritating effect on the skin due to the bristles (Ardelean and Mohan, 2008; Ciocârlan, 2009). The Galii aparinae herba contains iridoid glycosides, such as: asperulozidic acid, asperulozid, monotropein, aucubine (Deliorman and al., 2001); phenolic compounds like: p-cumaric and ferulic acids, flavonoids: luteoline, quercetine, rutoside, isoquercitrine and hyperoside and coumarins (Deliorman and al., 2001; Vlase et al., 2014); antraquinone derivatives of type nor-damnacant (Masanori et al., 2002). The roots present a red tannic substance, and the fruits contain caffeine, being used as a coffee substitute with lower caffeine content (Deliorman and al., 2001; Gruenwald et al., 2004). The plant also contains other alkaloids: protopine, harmine and hydroxydeoxypeganine derivatives (Gruenwald et al., 2004). The cleavers has been used since antiquity for healing wounds, burns, to eliminate the toxins from punctures and bites, or as a lymphatic drainer. The flowering herbs are used in varicos ulcer, skin irritations, but also in urinary lithiasis or as a diuretic (Gruenwald et al., 2004). The methanol extract from herba has been shown to be very effective on methicillin-resistant Staphylococcus aureus strains, which could be due to its rich content in polyphenolic compounds (flavonoids, phenolic acids, etc.) (Sharifi-Rad et al., 2016). The selective antiproliferative activity was shown on cellular lines of human breast cancer, noting that the methanol extract from G. aparine does not attack the normal epithelial cells (Atmaca et al., 2016). Another study highlighted the fact that the aqueous extract of Galium 85 aparine is a rich source of antioxidants, due to their polyphenolic content (Bokhari et al., 2013). Galium odoratum (L.) Scop., by its vernicular name woodruff, is a grassy, perennial species, widespread in Europe, from Spain to Ireland and Russia, but also in Asia, Turkey, Iran, from Caucasus to China and Japan. It has a pleasant smell due to the content in coumarins, which has also led to the cultivation of this species for ornamental purposes (Ardelean and Mohan, 2008; Ciocârlan, 2009). The plant has a height up to 30-50 cm and has more repent stem. It is very easy to be confused with Galium triflorum Michx. or Galium verum L. The simple, unhaired, lanceolate leaves are arranged 6-9 in whorls. The flowers are small, 4-7 mm in diameter, forming cyme, being white with 4 petals united at the base. The fruits are small (Ardelean and Mohan, 2008; Ciocârlan, 2009; Gruenwald et al., 2004). The woodruff fresh herb contains melilotoside, which during the drying is transformed into coumarin, whose concentration can reach up to 1%. It also contains iridoids, specific to the plant family: asperuloside, monotropeine, scandoside, aucubin (Gruenwald et al., 2004; Wojnicz et al. 2012). Beside the polyphenols were identified flavonoids, like apigenin derivatives, quercetine derivatives (e.g. rutoside, quercitrine) and phenolic acids, like p-coumaric acid, ferulic acid and cafeoilquinic acid derivatives (Vlase et al., 2014; Gruenwald et al., 2004). Due to the rich content in coumarins, Galium odoratum has anti- inflammatory, anti-edematous, lymphokinetic effects. The plant is indicated in nervous, hepatic, venous disorders (Gruenwald et al., 2004). An extensive study showed that the effect of this species is moderated on E. coli, but changes the hydrophobicity of the cellular surface, without inhibiting the motility of the bacterium, which loses its haemagglutination capacity (Vlase et al., 2014; Wojnicz et al. 2012). The water extract has a significant antioxidant effect, good healing effect in case of burns (Kahkeshani et al., 2013) and strong antibacterial activity on Staphylococcus aureus and Listeria monocytogenes (Vlase et al., 2014). Galium verum L., or lady’s bedstraw, is a perennial grassy plant, with a wide spread in Europe, North Africa and Asia, from Israel and Turkey to Japan and the Kamchatka Peninsula. It is considered an invasive plant. The stem grows up to 60-120 cm, the leaves are 1-3 cm long and 2 mm wide, dark green, hairy on the lower surface, forming whorls from 8-12 leaves. The flowers are small, yellow, in dense inflorescences (Ardelean and Mohan, 2008; Ciocârlan, 2009). Due to the smell given by the coumarins is used as repellent. In Scotland it is used for the clotting of milk, respectively the coloring of 86 cheese, containing a yellow dye. A specific lady’s bedstraw-based spirit drink is prepared in Denmark and the Scandinavian peoples are using it for the sedative effect (Howard, 1987). The lady’s bedstraw contains iridoids (asperuloside, monotropeine, scandoside, asperulosidic acid, etc.), flavonoids (rutoside, isorutoside, palustroside, cynaroside, isoquercitrine, quercitrine, quercetol, kaempferol, diosmetine), phenolic acids (chlorogenic acid, p-coumaric acid, caffeic acid, ferulic acid), catehic tannins, antraquinone derivatives (1.3-dihydroxy-2- methylantraquinone, fiscione, 2-hydroxy-1.3-dimethoxyantraquinone, 2.5- dihydroxy-1.3-dimethoxyantraquinone) and triterpenic acids (ursolic acid) (Vlase et al., 2014; Gruenwald et al., 2004; Mitova et al., 2002; Mavi et al., 2004; Zhao et al., 2006; Zhao et al., 2008). In the ethnopharmacological tradition the lady’s bedstraw tea is used as a sedative. Recent studies have highlighted that extracts of Galium verum exhibit potent anticandidiasis effect on different strains of Candida albicans, C. tropicalis and C. glabrata (Vlase et al., 2014; Yigit et al., 2009). Their extracts rich in flavonoids had anticancer, antioxidant effects and endothelial protector activity by increasing nitric oxide levels and endothelin secretion, inhibiting the thromboxanes and interleukins secretion (Schmidt et al., 2014; Zhao et al. 2011). Their antioxidant effect is due to inhibiting peroxidation and a potent inhibitor effect on Listeria monocytogenes was reveal (Vlase et al., 2014; Mavi et al., 2004). Another study revealed that the extract of G. verum has strong antihaemolitic effect, which is correlated with the powerful antioxidant effect (Khalili et al., 2014). Pharmacological studies have shown that the hydroalcoholic extract has capacity to activate the thyroid and ovaries in the case of mice subjected to anakinetic stress (Roman and Puică, 2013). The aim of this paper