This research was aimed to optimize the extraction conditions of three phenolic compounds: total phenolics, flavonoids and condensed tannins, from White Horehound's leaves (Marrubium vulgare L.). Distilled water and different organic solvents such as: methanol, ethanol and acetone, were used, with various concentrations (20% -80%, v/v), temperatures (20˚C -60˚C) and extraction times (30 -450 min). Results showed that the maximum total phenolics amounts (293.34 ± 14.60 mg gallic acid equivalent/g dry weigh), were obtained with 60% aqueous methanol at 25˚C for 180 min; total flavonoids (79.52 ± 0.55 mg catechin equivalent/g dry weigh) with 80% aqueous methanol at 20˚C for 450 min, and condensed tannins (28.15 ± 0.80 mg catechin equivalent/g dry weigh) with 60% aqueous acetone at 50˚C and for 180 min. ANOVA test showed the significant effect ( *** P < 0.001) of the extraction conditions tested on phenolic compounds. The Principal Component Analysis (PCA) exhibited the positive effect of low temperatures on total phenolics and flavonoids extraction, and the effect of high temperatures on the condensed tannins extraction. The Response Surface Methodology (RSM) provided predicted values of extraction conditions and maximum polyphenols amounts similar to those obtained experimentally.
IntroductionPolyphenols constitute one of the most common and widespread groups of substances in flowering plants, occurring in all vegetative organs, as well as in flowers and fruits. These molecules are involved in many physiological processes such as cell growth, root formation, seed germination and fruit ripening [1]. Moreover, these compounds are considered secondary metabolites involved in the chemical defense of plants against predators, pathogens, environmental stresses and in plant-plant interferences [2].Nowadays, phenolic compounds represent a unique and a functional place, composed of bioactive products, present in plant-derived foods and beverages and included in the formulations of well-marketed cosmetic and parapharmaceutical products [3]. Furthermore, polyphenols exhibit various biological activities such as anticancer [4], antioxidant [5], antimicrobial [6] and anti-inflammatory activities [7]. Therefore, in recent years, the determination of phenolic compounds concentrations in fruits [8] [9], vegetables [10] and some aromatic and medicinal plants [11] [12] has been of increasing interest in the scientific community as well as among health professionals and business partners. It is well known that the content of phenolic compounds could be influenced by environmental conditions, such as season [13] [14], sampling period and geographic origin [15], precipitations and temperatures [16], and soil type [17]. Additionally, there are several experimental factors that can influence the rate of extraction and the quality of extracted bioactive phenolic compounds. These factors include extraction method, solvent type used and concentration [18], particle size of medicinal plants, temperature and pH of extraction [19], extraction t...