Aspetti farmacologici e biotecnologici della produzione di alcaloidi da colture di cellule e tessuti di piante della famiglia delle amaryllidaceae

In this work particular attention was given to the study of secondary metabolites produced by some plants belonging to the Amaryllidaceae family, in the specific case isoquinoline alkaloids. At the first instance were characterized both qualitatively and quantitatively three different plants belonging to Amaryllidaceae family, such as: Crinum angustum Steud., Pancratium illyricum L., and Leucojum nicaeense Ard. The alkaloids extracts obtained were separately tested against enzymes involved in specific diseases or liable in multifactorial pathologies, like: MMPs, AChE,and PPO. From leaves extract of P.illyricum was isolated a new compound, 11α-hydroxy-O-methylleucotamine, with important role in AChE inbition. Considering the protection role against external bodies carried out by these metabolites in plant, extracts were also assayed against ATCC microorganisms and clinical isolates. Plants with promising pharmacological activities have been the basis for development of in vitro plant models

Mucuna pruriens in Parkinson Disease: A Kinetic-Dynamic Comparison With Levodopa Standard Formulations

Objectives: We compared levodopa (LD) kinetic-dynamic profile of a dose of LD/aromatic amino acid decarboxylase peripheral inhibitors versus a nominally equivalent dose of a commercial Mucuna pruriens (Mucuna) seeds extract in 2 patients with Parkinson disease chronically taking LD standard combined with self-prescribed Mucuna. Methods: Patients were challenged with a fasting morning dose of 100 mg LD/25 mg carbidopa (patient 1) or benserazide (patient 2) versus 100 mg LD from Mucuna capsules in 2 different sessions, after a 12-hour standard LD formulations\u2019 washout. They underwent kinetic-dynamic LD monitoring based on LD dose intake and simultaneous serial assessments of plasma drug concentrations and motor test performances. Quantitative analysis of LD in Mucuna capsules was also performed. Results: Levodopa bioavailability was markedly lower after Mucuna administration compared with LD standard formulations: in patient 1, peak plasma LD concentration (Cmax) decreased from 2.0 to 1.0 mg/L and the area under the plasma concentration time curve from 137 to 33.6 mg/L per minute; in patient 2, Cmax was 0.7 mg/L after LD/benserazide and nearly undetectable afterMucuna. In patient 1, impaired LD bioavailability from Mucuna resulted in reduced duration and overall extent of drug response compared with LD/carbidopa. In patient 2, no significant subacute LD motor response was observed in either condition. Quantitative analysis of Mucuna formulation confirmed the 100 mg LD content for the utilized capsules. Conclusions: Our results show an impaired LD bioavailability from Mucuna preparation, as expected by the lacking aromatic amino acid decarboxylase inhibitors coadministration,whichmight explain the suggested lower dyskinetic potential of Mucuna compared with standard LD formulations

Simultaneous analysis of diazepam and its metabolites in rat plasma and brain tissue by HPLC-UV and SPE

Diazepam is frequently used as an adjuvant during antidepressant therapy. Recently, some studies have suggested that the treatment with benzodiazepines could have different efficacy in depressed patients as opposed to non-depressed ones. To clarify the matter, a study is currently underway, regarding the drug metabolism in rats. In order to obtain a more complete and significant set of data, the main diazepam metabolites have also been considered, namely: nordiazepam, temazepam and oxazepam. A feasible and reliable HPLC method has been developed for the simultaneous determination of these compounds in plasma and brain tissue of rats. The method has been applied to "normal" rats and to genetic rat models of depression in order to estimate drug metabolism in different breeds. Analyte separation was achieved on a C8 reversed phase column using an acidic phosphate buffer/acetonitrile mixture as the mobile phase. The detection wavelength was 238 nm. An original sample pre-treatment, based on solid-phase extraction (SPE) was developed in order to eliminate endogenous interference, using only 250 μL of matrix (brain homogenate or plasma) for a complete analysis. The method has been validated with good results in terms of precision, extraction yield, sensitivity, selectivity and accuracy on both matrices and has been successfully applied to samples from some rats subjected to the preliminary study. The obtained data will hopefully contribute to the clarification of possible differences between depressed and non-depressed subjects with respect to benzodiazepine biotransformation. © 2009 Elsevier B.V. All rights reserved

Chemical composition, and in vitro antibacterial and antifungal activity of an alkaloid extract from Crinum angustum Steud.

In the search for novel antibacterial agents against multidrug-resistant bacteria, an alkaloid extract obtained from whole plants of Crinum angustum Steud., containing six different groups of alkaloids, was analysed by using gas chromatography/mass spectrometry, and its in vitro activity against American Type Culture Collection bacterial strains and clinical isolates was evaluated. The antimicrobial activity of the extract was tested against nine standard strains of microorganisms and two drugresistant clinical isolates, methicillin-resistant Staphylococcus aureus and carbapenemase- producing Klebsiella pneumoniae. The extract exhibited a significant activity against six of the examined strains of microorganisms. Antibacterial activity was more pronounced on Gram-positive bacteria than on Gram-negative bacteria, and IC50 values ranged from 156 to 625 mg/mL. Mycostatic activity against Candida albicans was also found, with an IC50 of 78 mg/mL after 48 h of incubation . Promising results were also obtained from tests carried out on both clinical isolates investigated

Dihydroasparagusic acid: Antioxidant and tyrosinase inhibitory activities and improved synthesis

Dihydroasparagusic acid (DHAA) is the reduced form of asparagusic acid, a sulfur-containing flavor component produced by Asparagus plants. In this work, DHAA was synthetically produced by modifying some published protocols, and the synthesized molecule was tested in several in vitro assays (DPPH, ABTS, FRAP-ferrozine, BCB, deoxyribose assays) to evaluate its radical scavenging activity. Results show that DHAA is endowed with a significant in vitro antioxidant activity, comparable to that of Trolox. DHAA was also evaluated for its inhibitory activity toward tyrosinase, an enzyme involved, among others, in melanogenesis and in browning processes of plant-derived foods. DHAA was shown to exert an inhibitory effect on tyrosinase activity, and the inhibitor kinetics, analyzed by a Lineweaver-Burk plot, exhibited a competitive mechanism. Taken together, these results suggest that DHAA may be considered as a potentially active molecule for use in various fields of application, such as pharmaceutical, cosmetics, agronomic and food. © 2013 American Chemical Society

Determination of phytomarkers in pharmaceutical preparations of Hemidesmus indicus roots by micellar electrokinetic chromatography and high-performance liquid chromatography-mass spectrometry

Micellar electrokinetic chromatography was applied to the determination of the major phytomarkers, namely 2-hydroxy-4-methoxybenzaldehyde, 2-hydroxy-4-methoxybenzoic acid, and 3-hydroxy-4-methoxybenzaldehyde, of Hemidesmus indicus root, an Indian medicinal plant. H. indicus bioactive preparations were analyzed by reverse flow micellar electrokinetic chromatography (MEKC) using sodiumtaurodeoxycholate as the surfactant. A pH 2.5 phosphate buffer (50mM) was supplemented with 65mM of sodium taurodeoxycholate to produce the MEKC pseudostationary phase; because of the suppression of the electroosmotic flow, the migration of the partitioned analytes was toward the capillary anodic end. The use of a short fused-silica capillary (8.5 cm effective length; 50 \ub5m i.d.) allowed the separation of phytomarkers, including vanillin and salicylaldehyde (reported as additional metabolites of H. indicus roots), in less than 8 min. The method showed good validation parameters and was applied to the analysis of methanol extracts and a root decoction of H. indicus, a promising botanical drug. The obtained results were compared to those from an independent high-performance liquid chromatography\u2013mass spectrometry method. The 2-Hydroxy-4-methoxybenzaldehyde, 2-hydroxy-4-methoxybenzoic acid, and 3-hydroxy 4-methoxybenzaldehyde were found in all samples confirming their roles as phytomarkers. The absence of vanillin and salicylaldehyde suggested that these latter compounds should not be regarded as characteristic components of the bioactive preparations from the plant roots

Dihydroasparagusic Acid: Antioxidant and Tyrosinase Inhibitory Activities and Improved Synthesis

Dihydroasparagusic acid (DHAA) is the reduced form of asparagusic acid, a sulfur-containing flavor component produced by <i>Asparagus</i> plants. In this work, DHAA was synthetically produced by modifying some published protocols, and the synthesized molecule was tested in several in vitro assays (DPPH, ABTS, FRAP-ferrozine, BCB, deoxyribose assays) to evaluate its radical scavenging activity. Results show that DHAA is endowed with a significant in vitro antioxidant activity, comparable to that of Trolox. DHAA was also evaluated for its inhibitory activity toward tyrosinase, an enzyme involved, among others, in melanogenesis and in browning processes of plant-derived foods. DHAA was shown to exert an inhibitory effect on tyrosinase activity, and the inhibitor kinetics, analyzed by a Lineweaver–Burk plot, exhibited a competitive mechanism. Taken together, these results suggest that DHAA may be considered as a potentially active molecule for use in various fields of application, such as pharmaceutical, cosmetics, agronomic and food