Anticancer activity of an Artemisia annua L. hydroalcoholic extract on canine osteosarcoma cell lines.

Since ancient times, Artemisia annua (A. annua) has been used as a medicinal plant in Traditional Chinese Medicine. In addition, recent studies have investigated the cytotoxic effects of A. annua extracts towards cancer cells. The leading aim of the present research is to evaluate the cytotoxic effects of an hydroalcoholic extract of A. annua on two canine osteosarcoma (OSA) cell lines, OSCA-8 and OSCA-40, focusing on the possible involvement of ferroptosis. The quantitative determination of artemisinin concentration in the extract, culture medium and OSA cells was carried out through the use of an instrumental analytical method based on liquid chromatography coupled with spectrophotometric detection and tandem mass spectrometry (LC-DAD-MS/MS). OSCA-8 and OSCA-40 were exposed to different dilutions of the extract for the EC50 calculation then the uptake of artemisinin by the cells, the effects on the cell cycle, the intracellular iron level, the cellular morphology and the lipid oxidation state were evaluated. A concentration of artemisinin of 63.8 ± 3.4 µg/mL was detected in the extract. A dose-dependent cytotoxic effect was evidenced. In OSCA-40 alterations of the cell cycle and a significantly higher intracellular iron content were observed. In both cell lines the treatment with the extract was associated with lipid peroxidation and with the appearance of a "ballooning" phenotype suggesting the activation of ferroptosis. In conclusion the A. annua idroalcoholic extract utilized in this study showed anticancer activity on canine OSA cell lines that could be useful in treating drug resistant canine OSAs.

Treatment of Benign Prostatic Hyperplasia by Natural Drugs.

Benign prostatic hyperplasia (BPH) is one of the most common urinary diseases affecting men, generally after the age of 50. The prevalence of this multifactorial disease increases with age. With aging, the plasma level of testosterone decreases, as well as the testosterone/estrogen ratio, resulting in increased estrogen activity, which may facilitate the hyperplasia of the prostate cells. Another theory focuses on dihydrotestosterone (DHT) and the activity of the enzyme 5α-reductase, which converts testosterone to DHT. In older men, the activity of this enzyme increases, leading to a decreased testosterone/DHT ratio. DHT may promote prostate cell growth, resulting in hyperplasia. Some medicinal plants and their compounds act by modulating this enzyme, and have the above-mentioned targets. This review focuses on herbal drugs that are most widely used in the treatment of BPH, including pumpkin seed, willow herb, tomato, maritime pine bark, Pygeum africanum bark, rye pollen, saw palmetto fruit, and nettle root, highlighting the latest results of preclinical and clinical studies, as well as safety issues. In addition, the pharmaceutical care and other therapeutic options of BPH, including pharmacotherapy and surgical options, are discussed, summarizing and comparing the advantages and disadvantages of each therapy.

Separation and non-separation methods for the analysis of cannabinoids in Cannabis sativa L.

Cannabis sativa L. is a plant known all over the world, due to its history, bioactivity and also social impact. It is chemically complex with an astonishing ability in the biosynthesis of many secondary metabolites belonging to different chemical classes. Among them, cannabinoids are the most investigated ones, given their pharmacological relevance. In order to monitor the composition of the plant material and ensure the efficacy and safety of its derived products, extraction and analysis of cannabinoids play a crucial role. In this context, in addition to a conventional separation method based on HPLC with UV/DAD detection, a new strategy based on a non-separation procedure, such as 13C-qNMR, may offer several advantages, such as reduced solvent consumption and simultaneous acquisition of the quali/quantitative data related to many analytes. In the light of all the above, the aim of this work is to compare the efficiency of the above-mentioned analytical techniques for the study of the main cannabinoids in different samples of cannabis inflorescences, belonging to fibre-type, recreational and medical varieties. The 13C-qNMR method here proposed for the first time for the quantification of both psychoactive and non-psychoactive cannabinoids in different cannabis varieties provided reliable results in comparison to the more common and consolidated HPLC technique.

Development of an electrochemical sensor based on carbon black for the detection of cannabidiol in vegetable extracts.

A glassy carbon electrode chemically modified with a carbon black coating is proposed here for the rapid and portable determination of cannabidiol (CBD) in a commercial Cannabis seed oil and in fibre-type Cannabis sativa L. leaves. The mechanism of CBD oxidation was studied in relation to simpler phenyl derivatives bearing the same electroactive group, namely resorcinol and 2-methylresorcinol. These molecules also allowed us to determine the best conditions for the electrochemical detection of CBD, as to the pH value and to the best solvent mixture to use. Carbon black was chosen among nanostructured carbon-based materials owing to its outstanding features as an electrode modifier for analyte detection. The performance of the modified electrode was determined by flow injection analyses of standard solutions of CBD, obtaining a linear correlation between the oxidation current and the analyte concentration; the sensor response is characterised by suitable repeatability and reproducibility. The analysis of commercial products by the standard addition method allowed us to ascertain the accuracy of the sensor for the detection of CBD in real samples.

Analysis of Artemisia annua extracts and related products by high performance liquid chromatography-tandem mass spectrometry coupled to sample treatment miniaturisation.

Artemisinin, the main antimalarial compound of Artemisia annua L., is currently attracting increasing interest for its antiproliferative properties, but its content is highly variable, depending on several genetic, environmental and processing conditions. Aim of the present study is to analyse the artemisinin content in different plant extracts, to test their in vitro activity on cell proliferation and then to correlate these data to the active principle concentration. For this purpose, an innovative miniaturised sample pretreatment strategy based on microextraction by packed sorbent (MEPS) was developed and coupled to an original advanced method based on liquid chromatography with diode array detection and tandem mass spectrometry (LC-DAD-MS/MS). The method was fully validated, granting consistent data. Good linearity was found over a suitable concentration range, i.e. 5-1000ng/mL. Extraction yields (>85%), precision (RSD < 3.5%) and accuracy (recovery 88-93%) were all within acceptable levels of confidence. After validation, the method was successfully applied to the determination of artemisinin in A. annua extracts. Analyte content was widely variable (up to twenty-fold) according to the starting material and the extraction procedure, ranging between 5.9µg/g and 109µg/mL. The cytotoxic activity of all analysed extracts was also tested on human leukemic cells by viable cell count and cell cycle analysis. Artemisinin concentrations and biological activity were carefully evaluated and the observed antiproliferative effects varied according to artemisinin content in each extract type. This highlights the need to quantitatively analyse the main active constituent of plant extracts and the obtained data have shown to be promising for the choice of the related herbal product dosage.

Encapsulations of wild garlic (Allium ursinum L.) extract using spray congealing technology.

The objective of this study was to incorporate wild garlic (A. ursinum) extract into microparticles (MPs) in order to protect its valuable active compounds and improve its oral bioavailability. For this purpose, spray congealing technology was applied and Gelucire 50/13 (Stearoyl polyoxyl-32 glycerides) was selected as MPs carrier. MPs were characterized in terms of yield, encapsulation efficiency and particle size. Differential scanning calorimetry (DSC) and Fourier-transform infrared spectroscopy (FT-IR) analysis of MPs showed the absence of chemical interactions between carrier and extract and suggested that spray congealing process did not modify nor degrade the encapsulated extract. The encapsulation into MPs led to an improvement of the extract dissolution performance as well as an enhancement in solubility of >18 fold compared to the pure extract. Additionally, MPs were stable over three months showing only a minor decrease in the content of active compounds (allicin and S-methyl methanethiosulfonate) and maintaining a good antimicrobial activity. Therefore, obtained results suggested that the encapsulation of A. ursinum extract in MPs by spray congealing is a promising approach to improve the biopharmaceutical properties of the extract, without affecting its antibacterial activity.

Analytical Profiling of Bioactive Phenolic Compounds in Argan (Argania spinosa) Leaves by Combined Microextraction by Packed Sorbent (MEPS) and LC-DAD-MS/MS.

INTRODUCTION: The argan tree (Argania spinosa) is an endemic species from south-western Morocco. Argan-based preparations have been widely used in Moroccan traditional medicine for their biological properties, as well as for several cosmetic purposes. Whereas kernel, pulp of fruit and trunk have been extensively studied for their nutritional and pharmacological effects, relatively little is known about argan tree leaves. OBJECTIVE: The main purpose of the present study is to investigate and characterise the bioactive phenolic fractions in both crude and aqueous extracts derived from argan tree leaves. METHODOLOGY: A qualitative profile of the antioxidant phenolic compounds in argan leaves was obtained by means of structural hypothesis based on UV spectra and mass spectrometric fragmentation patterns. Moreover, selected phenolics were quantified in argan leaves by using a fully validated method based on liquid chromatography coupled to diode array detection and tandem mass spectrometry (LC-DAD-MS/MS). All the extracts were purified by a fast and reliable microextraction by packed sorbent (MEPS) procedure, before analysing them by LC-MS/MS. RESULTS: Based on retention times, mass spectrometric fragmentation and UV spectra, 13 phenolic compounds were identified or tentatively elucidated from crude and aqueous extracts derived from Argania spinosa leaves, while seven compounds were quantified in both extracts. CONCLUSION: The obtained results could represent a first step towards a complete characterisation of the argan plant, its bioactive profiling and the valorisation of its by-products as a source of potentially beneficial bioactive molecules.

Quantitative evaluation of auraptene and umbelliferone, chemopreventive coumarins in citrus fruits, by HPLC-UV-FL-MS.

An analytical strategy, based on the development of two HPLC methods with spectrophotometric (UV), spectrofluorometric (FL), and mass spectrometric (MS) detection, has been developed to investigate the presence of and to quantitate two important chemopreventive coumarins, auraptene and umbelliferone, in foodstuffs. The analytes were determined in fruits, and fruit parts, of plants belonging to the Citrus , Poncirus , and Fortunella genera, to test their nutraceutical potential. The method validation has been carried out according to international guidelines, with good results in terms of precision (RSD < 6.9%) and extraction yields (>91%). Application to the quantitative analysis of auraptene and umbelliferone in several kinds of citrus fruits was successful, providing reliable and consistent data. Exploiting three different kinds of detection, the analytical methodology proposed herein has been demonstrated to be sound but versatile, as well as reliable. Performances and results were compared and always found in good agreement among themselves. Thus, this approach is suitable for the identification and simultaneous quantitation of auraptene and umbelliferone in citrus fruits, with the aim of evaluating their nutraceutical potential.