Alfredo Aires

The antimicrobial effects of glucosinolates and their respective enzymatic hydrolysis products on bacteria isolated from the human intestinal tract

Aims:  The aim of the study was to evaluate the in vitro antibacterial activity of glucosinolates and their enzymatic hydrolysis product against bacteria isolated from the human intestinal tract.

Antimicrobial activity of phenolics and glucosinolate hydrolysis products and their synergy with streptomycin against pathogenic bacteria.

The purpose of the present study was to evaluate the in vitro antibacterial effects of different classes of important and common dietary phytochemicals (5 simple phenolics - tyrosol, gallic acid, caffeic acid, ferulic acid, and chlorogenic acid; chalcone - phloridzin; flavan-3-ol - (-) epicatechin; seco-iridoid - oleuropein glucoside; 3 glucosinolate hydrolysis products - allylisothiocyanate, benzylisothiocyanate and 2-phenylethylisothiocyanate) against Escherichia coli, Pseudomonas aeruginosa, Listeria monocytogenes and Staphylococcus aureus. Another objective of this study was to evaluate the effects of dual combinations of streptomycin with the different phytochemicals on antibacterial activity. A disc diffusion assay was used to evaluate the antibacterial activity of the phytochemicals and 3 standard antibiotics (ciprofloxacin, gentamicin and streptomycin) against the four bacteria. The antimicrobial activity of single compounds and dual combinations (streptomycin-phytochemicals) were quantitatively assessed by measuring the inhibitory halos. The results showed that all of the isothiocyanates had significant antimicrobial activities, while the phenolics were much less efficient. No antimicrobial activity was observed with phloridzin. In general P. aeruginosa was the most sensitive microorganism and L. monocytogenes the most resistant. The application of dual combinations demonstrated synergy between streptomycin and gallic acid, ferulic acid, chlorogenic acid, allylisothiocyanate and 2-phenylethylisothiocyanate against the Gram-negative bacteria. In conclusion, phytochemical products and more specifically the isothiocyanates were effective inhibitors of the in vitro growth of the Gram-negative and Gram-positive pathogenic bacteria. Moreover, they can act synergistically with less efficient antibiotics to control bacterial growth.

Seasonal effects on bioactive compounds and antioxidant capacity of six economically important brassica vegetables.

Research on natural and bioactive compounds is increasingly focused on their effects on human health, but there are unexpectedly few studies evaluating the relationship between climate and natural antioxidants. The aim of this study was analyze the biological role of six different Brassica vegetables (Brassica oleracea L. and Brassica rapa L.) as a natural source of antioxidant compounds. The antioxidant activity may be assigned to high levels of L-ascorbic acid, total phenolics and total flavonoids of each sample. The climate seasons affected directly the concentration of bioactive components and the antioxidant activity. Broccoli inflorescences and Portuguese kale showed high antioxidant activity in Spring-Summer whilst turnip leaves did so in Summer-Winter. The Brassica vegetables can provide considerable amounts of bioactive compounds and thus may constitute an important natural source of dietary antioxidants.

Initial in vitro evaluations of the antibacterial activities of glucosinolate enzymatic hydrolysis products against plant pathogenic bacteria.

Aims:  The aim of the study was to evaluate the in vitro antibacterial effects of glucosinolate hydrolysis products (GHP) against plant pathogenic micro‐organisms namely Agrobacterium tumefaciens, Erwinia chrysanthemi, Pseudomonas cichorii, Pseudomonas tomato, Xanthomonas campestris and Xanthomonas juglandis.

Genetic organisation of Iris yellow spot virus M RNA: indications for functional homology between the G(C) glycoproteins of tospoviruses and animal-infecting bunyaviruses

Summary. The complete nucleotide sequence (4838 nucleotides) of Iris yellow spot virus (IYSV) M RNA indicates, typical for tospoviruses, the presence of two genes in ambisense arrangement. The vRNA ORF codes for the potential cell-to-cell movement (NSm) protein (34.8 kDa) and the vcRNA ORF for the viral glycoprotein (G1/G2) precursor (128.6 kDa). Multiple sequence alignment of the NSm and G1/G2 precursor proteins of IYSV with those of other tospoviruses, showed highest homologies to Peanut bud necrosis virus (PBNV) and Watermelon silver mottle virus (WSMV). The potential cell-to-cell movement protein of tospoviruses is highly conserved (40–70% identity), with the exception of the first 60 N terminal amino acids, a domain that clearly diverged. For the G1 and G2 viral glycoproteins, blast searches revealed a significant homology between the C-terminally located tospoviral G1 (G(C)) protein with the counterpart of the animal-infecting bunyaviruses, suggesting a functional homology for these proteins.

Phytochemical characterization and antioxidant properties of baby-leaf watercress produced under organic production system

The aim of this study was to evaluate the phytochemical composition and the antioxidant properties of watercress (Nasturtium officinale R. Br., Brassicaceae) produced under organic production system. Fresh baby-leaf watercress samples were collected from local organic farms. High performance liquid chromatography with diode array detection (HPLC-DAD) and high performance liquid chromatography with diode array detection mass spectrometry (HPLC-MS) were used to assess the phytochemical composition, and spectrophotometric methods were used to assess the antioxidant capacity. Two major classes of healthier secondary plant metabolites were identified: phenolics and glucosinolates. The major phenolics determined were chlorogenic acid, quercetin-3-O-rutinoside, dicaffeoyltartaric acid, and isorhamnetin. The glucosinolates were composed exclusively of gluconasturtiin, the precursor of the anticarcinogenic, and antimicrobial compound 2-phenylethyl isothiocyanate. The extracts of organic young baby-leaf watercress presented high antioxidant capacity and this property was highly related with caffeic acid, quercetin-3-O-rutinoside, isorhamnetin, and glucosnastrutiin. The results achieved showed that baby-leaf watercress can provide high contents of antioxidant compounds at levels even superior to similar adult plant material.

Antibacterial activity and synergistic effect between watercress extracts, 2-phenylethyl isothiocyanate and antibiotics against 11 isolates of Escherichia coli from clinical and animal source

To evaluate the possible in vitro interaction between natural extracts of watercress (Nasturtium officinale R. Br.) and 2‐phenylethyl isothiocyanate, a natural compound derived gluconasturtiin largely present in watercress tissues, with a standard antibiotic, a synergy study was carried out against 11 isolates of extended‐spectrum β‐lactamases‐Escherichia coli. Aqueous and methanolic watercress extracts and 2‐phenylethyl isothiocyanate were combined with the antibiotic, and a disc diffusion assay and minimum inhibitory concentration methods were used to assess the in vitro antibacterial activity. The results of this study showed that there is an increase in antibacterial activity of the antibiotic when it was combined with plants extracts and pure compounds. The most interesting result was the combination between 2‐phenylethyl isothiocyanate and the antibiotic. Synergistic effects of the antibiotic with watercress extracts and 2‐phenylethyl isothiocyanate suggest the potential of these plants and their natural compounds to improve the performance of the antibiotics and could be an interesting tool for antimicrobial therapy. The results led us to conclude that watercress has important pharmacological substances which can be used for developing new and effective antimicrobial agents.

Glucosinolate assessment in Brassica oleracea leaves by near-infrared spectroscopy

Glucosinolates and their hydrolysis products are of great concern because they are responsible for many of both the beneficial and harmful properties of glucosinolate-containing plants. Cabbage ( Brassica oleracea L.) is a species highly consumed as a leaf vegetable in many countries showing high qualitative and quantitative differences in glucosinolate composition among cultivars. The standard analytical techniques for determining glucosinolate composition using conventional methods lead to high costs, labour input and delays, all of which affect both the availability of data and the taking of swift decisions. In contrast, near-infrared spectroscopy (NIRS) has emerged as a rapid and cost-effective technique of analysis for many agro-food products. In this work we test the potential of NIRS for screening the total glucosinolates (t-GSL), gluconapin (GNA), gluconasturtiin (GNAST) and neoglucobrassicin (NGBS) contents of cabbage leaf cultivars coming from Portugal and Spain. NIRS calibrations resulted in coefficients of determination and standard deviation to standard error of cross-validation ratio of 0·83 and 2·38 for t-GSL; 0·70 and 1·85 for GNA; 0·62 and 1·63 for GNAST; and 0·60 and 1·58 for NGBS, respectively. An examination of the loadings of the equation for t-GSL suggested that O-H groups of water, C-H combinations of the methylene group and also N-H groups of amides were the molecular associations most strongly used in modelling total glucosinolates. It was concluded that NIRS shows a high potential as an analytical method for total and individual glucosinolate routine analysis in cabbage.

Antimicrobial Activity of Isothiocyanates from Cruciferous Plants against Methicillin-Resistant Staphylococcus aureus (MRSA)

Purified isothiocyanates from cruciferous plants (Brassicacea, Syn. Cruciferae) plants were evaluated against 15 isolates of methicillin-resistant S. aureus isolated from diabetic foot-ulcer patients aiming the study of the potential usage of allyl-isothiocyanate, benzyl-isothiocyanate and 2-phenylethyl-isothiocyanate against this important bacteria. Disc diffusion and minimum inhibitory concentration methods were used to access the antimicrobial activity. The index (Ia) and rate (Ra) of the antibacterial activity for each compound were calculated. The results showed a highly dose-dependent compound and chemical structure antibacterial effectiveness. The results showed a strong relation between the chemical structure of isothiocyanates and its antibacterial effectiveness. The benzyl-isothiocyanate was the most effective with a minimum inhibitory concentration varying between 2.9 and 110 µg· mL−1 with an antibacterial activity rate up to 87%. Moreover, their antibacterial activity was mainly bactericidal. This study provides scientific evidence that isothiocyanates have an interesting biological value and must be considered as an important tool to be used against MRSA.

Influence of Nitrogen and Sulfur Fertilization on the Mineral Composition of Broccoli Sprouts

ABSTRACT Broccoli sprouts (Brassica oleraceae var. italica) have been attributed health protective effects based on their glucosinolate content, and thus, are recommended in diets. However, no information is available on the mineral content of this novel product and how fertilization might influence it. The influence of nitrogen (N) and sulfur (S) applications (0, 14, and 28 mg· N dish−1 and 0, 4.5, and 9 mg· S dish−1) on the mineral content [N, S, potassium (K), calcium (Ca), magnesium (Mg), sodium (Na), chloride (Cl), and silicon (Si)] of broccoli sprouts (Brassica oleraceae var. italica cv. ‘Marathon’) was determined 11 d after sowing. It was found that N and S fertilization significantly (P < 0.001) influenced the uptake of all elements except phosphorus (P). Sulfur concentrations in broccoli sprouts varied between 11.4 and 15.2 mg· g−1 (dw), while the Ca, Mg, P, K, and Na concentrations were below 10 mg· g−1 (dw). The Cl contents ranged from 13.6 to 23.1 mg· g−1 (dw). The highest S concentration was found when 9 mg· dish−1 S and 14 mg· dish−1 N was applied. A higher N rates of 28 mg· dish−1, N yielded no higher S uptake. The significantly (P < 0.05) highest Ca, Mg, and Na concentrations were found in the control treatments, while this effect proved to be not consistent for P. The results clearly revealed that N and S fertilization increased biomass production even in the early growth stages. With view to maintaining high Ca concentrations an application in the form of ammonium sulfate would be preferable.