Study of chemical composition and biological activities of Euphorbia resinifera and Euphorbia officinarum and their honeys

Boutoub, Oumaima

http://hdl.handle.net/10400.1/26959

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Authors

Boutoub, Oumaima

Advisor(s)

Miguel, Maria da Graça Costa

Abstract(s)

Euphorbia officinarum and Euphorbia resinifera are two species found in Morocco. They have been used in traditional medicine. Reports indicate the use of Euphorbia officinarum for the treatment of wounds, skin infections and abscesses and the aerial parts of Euphorbia resinifera in combination with honey or extracts obtained by decoction have been used by patients in Morocco to treat cancer. The “Morocco Green Plan” has played an important in terms of sustainable development of the agricultural sector, helping to increase its benefit, through the promotion of specific products from different territories. One of these products is honey. Propolis (a natural substance produced by bees from plant resins, sap and other botanical sources) is beginning to attract the interest of Moroccan beekeepers thanks to its biological properties. This work does not focus on latex, but on the aerial parts of E. officinarum and E. resinifera, the inflorescence part called cyathium of E. resinifera, which do not secrete latex; monofloral honey of E. officinarum and E. resinifera; as well as propolis from E. officinarum and E. resinifera. The solvent for extracting the aerial parts of both species was water, because it is the liquid used by the population to treat their illnesses. The extraction temperature; extraction time; and plant/solvent ratio are the parameters chosen to obtain extracts with the highest quantities of phenols and with the best in vitro antioxidant properties, chelating activity and capacity to inhibit α-glucosidase activity using response surface methodology (RSM). The best extraction conditions detected were using 1 g/50 mL during 60 minutes at 30 ºC for E. resinifera and for E. officinarum; the best conditions were temperature of 50 ºC, using1 g/100 mL for 270 minutes. Euphorbia resinifera cyathium releases a nectar that bees collect and transform into honey, currently labeled as a Protected Geographical Indication, due to its qualities. Additionally, flower cyathium honey and water extracts have been used in traditional medicine in Morocco. Several compounds belonging to different metabolite classes were identified in the present investigation. Concerning the primary metabolites: in hexoses, glucose followed by fructose predominates; in organic acids, citrate and α-ketoglutarate were the most abundant; amino acids were also found in abundance, notably asparagine (Asn); and vitamins such as niacin, vitamin B3, threonate (a metabolite of vitamin C), and the antioxidants ascorbate and dehydroascorbate. Regarding secondary metabolites: lutein was dominant in the pigments identified; flavonoids were dominated by the flavonol glycosides quercetin, kaempferol and isorhamnetin; polyamines were also identified. The scavenging capacity of DPPH and superoxide radicals by the floral extracts corresponded to two thirds and half of the capacity of pure compounds used respectively as positive control. Out of seven honey samples, two were of floral origin from E. resinifera and after pollen analysis, the other five were found to have high percentages of E. officinarum pollen grains. The potassium (K) was the most important mineral element in in all samples with an average content of 409 mg/kg. The chromatographic profiles evaluated by liquid chromatography coupled to diode array detection and mass spectrometry (LC-DAD-MS) of Euphorbia honey extracts were quite different. In all samples, it was possible to detect gallic acid, 4-hydroxybenzoic acid and p-coumaric acid, although in different proportions. Naringenin was identified in all but one sample. Abscisic acid was detected in five of seven honey samples, being the major compound detected in in one E. resinifera honey. The antioxidant properties of E. officinarum and E. resinifera as well as their ability to inhibit acetylcholinesterase, lipoxygenase, tyrosinase, and xanthine oxidase enzymes were evaluated and compared. At the same time, the in vitro biological activities of the entire honey and the extracts obtained from the respective honey were also evaluated and compared. Generally, the methanolic extracts of the honey recorded better activities in all the biological activities, than the corresponding entire honey. The antioxidant properties and the capacity to inhibit some enzymatic activities of E. resinifera and E. officinarum aqueous extracts and honeys were assessed. The aqueous extracts were obtained by decoction and two extraction parameters were chosen: extraction time (1 h, 2 h, and 8 h); and PSR (plant solvent ratio) of 1 g/20 mL; 1 g/50 mL, and 1 g/100 mL. The physicochemical characteristics of the honey were also evaluated and were within the range of the quality standards, E. officinarum honey showed higher amounts of total phenols and better capacity for scavenging superoxide anion free radicals and DPPH (2,2-diphenyl-1-picrylhydrazyl) free radicals than E. resinifera honey, but poorer capacity for inhibiting lipoxygenase, acetylcholinesterase, tyrosinase, and xanthineoxidase. Moreover, the aqueous extracts had systematically higher in vitro activities than the respective honey samples. Concerning propolis samples, the study aimed to characterize the pollen type, the volatile compounds, the phenolic and mineral profiles of three Euphorbia propolis samples collected in Morocco. In addition, it was intended to evaluate its antioxidant and antimicrobial activities, as well has the enzymatic inhibition potential. Pollen analysis revealed that E. resinifera pollen dominated in the P1 sample (58%), while E. officinarum pollen dominated in the P2 and P3 samples (44%). The volatile compounds were primarily composed of monoterpenes hydrocarbons, with α-pinene being the major component. Calcium (Ca) was the predominant mineral element in all propolis samples. Higher levels of phenols, flavonoids, and dihydroflavonoids were detected in the E. officinarum which corresponded to superior antioxidant activity. This was determined by its ability to scavenge DPPH and nitric oxide (NO) free radicals, superoxide anion radicals, and to prevent lipid peroxidation. P1 and P3 exhibited the best inhibitory effects on glucosidase activity, while the P2 sample demonstrated the highest efficacy in inhibiting acetylcholinesterase, lipoxygenase, tyrosinase, and xanthine oxidase activities. Euphorbia propolis displayed the ability to inhibit quorum sensing in the biosensor Chromobacterium violaceum CV026 and disrupted bacterial biofilm formation, including that of resistant bacterial pathogens.

Euphorbia officinarum e E. resinifera são duas espécies que se podem encontrar em Marrocos. Têm sido usadas em medicina popular. Relatos indicam a utilização de E. officinarum para o tratamento de feridas, infecções cutâneas e abcessos ou, em associação com outras espécies locais (Ziziphus lotus and Opuntia ficus-barbarica) ou com mel, no tratamento de pielonefrite ou nefrite. As partes aéreas de E. resinifera em associação com mel ou extractos obtidos por decoccão têm sido usadas, por doentes de Marrocos, no tratamento do cancro, enquanto que o látex fresco é usado em picadas venenosas, mordeduras e dores de dentes. O “Plan Vert Marroc” tem sido de grande importância no que ao desenvolvimento sustentável do sector agrícola diz respeito, contribuindo para aumentar o seu valor acrescido, mediante a promoção dos produtos específicos dos diversos territórios com a indicação de Indicações Geográficas, Designação de Origem ou Rótulos Agrícolas. Um desses produtos é o mel. Dois méis de origem monofloral de Euphorbia foram registados, designadamente: indicação geográfica “mel do deserto Euphorbia” e indicação de protecção geográfica “mel de Euphorbia de Tadla-Azilal”. A espécie da planta em concreto não foi providenciada, contudo é possível encontrar três tipos de mel monofloral em Marrocos: Euphorbia officinarum subsp. echinus, E. regis-jubae, and E. resinifera. E. resinifera. Própolis (uma substância natural produzida pelas abelhas a partir de resinas de plantas, seiva e outras fontes botânicas) começa a suscitar o interesse dos apicultores marroquinos graças às suas propriedades biológicas. A nível mundial, o própolis tem sido genericamente usado na medicina tradicional ou em suplementos alimentares. Desta forma, este produto apícola pode constituir uma fonte adicional de receitas para os apicultores marroquinos. A revisão bibliográfica efectuada sobre E. officinarum e E. resinifera demonstrou que o foco da investigação científica tem sido o látex. Tem existido grande interesse por E. officinarum para a obtenção de compostos por hemissíntese a partir de compostos isolados do látex com o objetivo de melhorar as atividades biológicas. A partir do látex de E. resinifera é possível isolar a resiniferatoxina (diterpeno extremamente irritante) que tem sido usado nalguns ensaios clínicos como um potencial analgésico no alívio de dor oncológica e artrítica, sendo cerca de 1000 vezes mais potente que a capsaicina. O enfoque deste trabalho incide não sobre o látex, mas sim sobre as partes aéreas de E. officinarum e E. resinifera sem o látex e sem as pseudo-inflorescências, designadas ciátio; ciátio de E. resinifera, que não secretam látex; mel monofloral de E. officinarum e E. resinifera; bem como o própolis de E. officinarum e E. resinifera.