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Development of a flow injection analytical system for short chain amide determination based on a tubular bioreactor and an ammonium sensor

Publication . Veríssimo, M.I.S.; Oliveira, Sara B.; Silva, Nelson A. F.; Matos, Manuel; Karmali, Amin; Gomes, Maria Teresa SR

Pseudomonas aeruginosa (P. aeruginosa) possesses intracellular amidase activity, which catalyses the hydrolysis of short aliphatic amides producing NH4 +, and has already been used along with an ammonium ion selective electrode for amide quantification. However, the incorporation of a biological membrane turned to be a challenging process and either the final arrangement was prone to amidase losses or the recovery of the sensor coating after the interaction took too long. In this article a flow injection system with an ammonium acoustic wave sensor is proposed, and after testing several different arrangements for the biological element, the ultimate choice consisted of the immobilization of a P. aeruginosa cell-free extract in the inner wall of a tubular glass reactor, which resulted in a reliable analytical system. Response times less than one minute and complete recovery in less than two minutes assured conveniently fast analysis. The analytical system, as long as the column was properly stored in HEPES buffer containing 2 mM β-mercaptoethanol and 1 mM benzamidine and refrigerated when not in use, could be used at least for 20 working days, along a period of one month, maintaining the initial sensitivity.

2018-08-21Journal article

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Bioactivities of decoctions from Plectranthus species related to their traditional use on the treatment of digestive problems and alcohol intoxication

Publication . Brito, Elsa; Gomes, Emma; Fale, Pedro; Borges, Carlos; Pacheco, Rita; Teixeira, Vítor; Machuqueiro, Miguel; Ascensão, Lia; Serralheiro, Maria Luisa

Ethnopharmacological relevance Decoctions of Plectranthus species are traditionally ingested after large meals for treatment of food digestion and alcohol abuse. Aim of the study This study aims at associating the digestion-related ethno-uses of Plectranthus species decoctions to molecular mechanism that might explain them: easing digestion (AChE inhibition) and treating hangover (ADH inhibition) Material and methods Decoctions from Plectranthus species were analysed for their alcohol dehydrogenase (ADH) inhibition and acetylcholinesterase (AChE) inhibition, related with alcohol metabolism and intestinal motility, respectively. Identification of the active components was carried out by LC-MS/MS and the docking studies were performed with AChE and the bioactive molecules detected. Results All decoctions inhibited ADH activity. This inhibition was correlated with their rosmarinic acid (RA) content, which showed an IC50 value of 19 μg/mL, similar to the reference inhibitor CuCl2. The presence of RA also leads to most decoctions showing AChE inhibiting capacity. P. zuluensis decoction with an IC50 of 80 μg/mL presented also medioresinol, an even better inhibitor of AChE, as indicated by molecular docking studies. Furthermore, all decoctions tested showed no toxicity towards two human cell lines, and a high capacity to quench free radicals (DPPH), which also play a helpful in the digestive process, related with their RA content. Conclusions All activities presented by the RA-rich Plectranthus decoctions support their use in treating digestion disorders and P. barbatus could explain its use also for alleviating hangover symptoms. Medioresinol, which is present in P. zuluensis, exhibited a significant AChE inhibition and may provide, in the future, a new lead for bioactive compounds.

2018-06-28Journal article

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Inhibition of HMG-CoA reductase activity and cholesterol permeation through Caco-2 cells by caffeoylquinic acids from Vernonia condensata leaves

Publication . Arantes, Ana A.; Fale, Pedro L.; Costa, Larissa C. B.; Pacheco, Rita; Ascensão, Lia; Serralheiro, Maria Luísa

The aim ofthis study was to provide scientific knowledge to supportthe use of Vernonia condensata Baker, Asteraceae, beverages for their alleged hypocholesterolemic properties by testing their action as HMGCoA reductase inhibitors and their capacity to lower dietary cholesterol permeation. Chlorogenic acid, and other caffeoylquinic acids derivatives were identified as the main components of these beverages by LC–MS/MS. No changes in the composition were notice after the in vitro gastrointestinal digestion and no toxicity against Caco-2 and HepG2 cell lines was detected. Cholesterol permeation through Caco-2 monolayers was reduced in 37% in the presence of these herbal teas, and the caffeoylquinic acids permeated the monolayers in 30–40% of their initial amount in 6 h. HMG-CoA reductase activity was reduced with these beverages, showing an IC50 of 217 g ml−1. It was concluded that caffeoylquinic acids, the major components, justified 98% of the enzyme inhibition measured.

2016Journal article

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