Response surface optimisation of extraction of antioxidants from strawberry fruit, and lipid peroxidation inhibitory potential of the fruit extract in cooked chicken patties.

BACKGROUND: Strawberries contain high levels of antioxidants and have beneficial effects against oxidative stress-mediated diseases, such as cancer. They contain multiple phenolic compounds, which contribute to their biological properties. Hence, a study was carried out to optimise the extraction of antioxidants and evaluate the antioxidant potential of strawberry fruit extract (SE) in cooked chicken patties during refrigerated storage. The activity of SE was compared with that of butylhydroxytoluene (BHT). RESULTS: The effect of solvent type (MeOH and EtOH), concentration (0-70%) of EtOH in the system, temperature (30-60 °C), and time (30-150 min) on DPPH•-scavenging activity of SE was investigated. Response surface methodology was used to estimate the optimum extraction conditions for each parameter. The maximum predicted DPPH• scavenging under the optimised conditions (100% MeOH, 30 °C, 150 min) was 43% at 1 mg SE mL⁻¹. Freshly prepared chicken patties were treated with 5% and 10% SE and 2% BHT, and stored aerobically at 4 °C for 6 days. SE had no influence (P < 0.05) on any of the sensory attributes of the patties. The values of thiobarbituric acid reactive substances reduced significantly (P < 0.05) from 2.47 mg in control patties to 0.312 mg and 0.432 mg malonaldehyde kg⁻¹ sample in 5-SE and 10-SE patties, respectively, on the day 6 of storage. CONCLUSION: The optimised model depicted MeOH at 30 °C with an extended time of 150 min as the optimum settings for extraction of compounds from strawberry that had the scavenging activity. The study shows that the extraction of natural antioxidants from strawberry can be improved by optimising several key extraction parameters. SE also acted as an effective antioxidant and suppressed lipid peroxidation in cooked chicken patties.

Novel non-peptidic vinylsulfones targeting the S2 and S3 subsites of parasite cysteine proteases.

We describe here the identification of non-peptidic vinylsulfones that inhibit parasite cysteine proteases in vitro and inhibit the growth of Trypanosoma brucei brucei parasites in culture. A high resolution (1.75 A) co-crystal structure of 8a bound to cruzain reveals how the non-peptidic P2/P3 moiety in such analogs bind the S2 and S3 subsites of the protease, effectively recapitulating important binding interactions present in more traditional peptide-based protease inhibitors and natural substrates.

Nonpeptidic tetrafluorophenoxymethyl ketone cruzain inhibitors as promising new leads for Chagas disease chemotherapy.

A century after discovering that the Trypanosoma cruzi parasite is the etiological agent of Chagas disease, treatment is still plagued by limited efficacy, toxicity, and the emergence of drug resistance. The development of inhibitors of the major T. cruzi cysteine protease, cruzain, has been demonstrated to be a promising drug discovery avenue for this neglected disease. Here we establish that a nonpeptidic tetrafluorophenoxymethyl ketone cruzain inhibitor substantially ameliorates symptoms of acute Chagas disease in a mouse model with no apparent toxicity. A high-resolution crystal structure confirmed the mode of inhibition and revealed key binding interactions of this novel inhibitor class. Subsequent structure-guided optimization then resulted in inhibitor analogues with improvements in potency despite minimal or no additions in molecular weight. Evaluation of the analogues in cell culture showed enhanced activity. These results suggest that nonpeptidic tetrafluorophenoxymethyl ketone cruzain inhibitors have the potential to fulfill the urgent need for improved Chagas disease chemotherapy.

Proteomic analysis of skin invasion by blood fluke larvae.

BACKGROUND: During invasion of human skin by schistosome blood fluke larvae (cercariae), a multicellular organism breaches the epidermis, basement membrane, and dermal barriers of skin. To better understand the pathobiology of this initial event in schistosome infection, a proteome analysis of human skin was carried out following invasion by cercariae of Schistosoma mansoni. METHODOLOGY AND RESULTS: Human skin samples were exposed to cercariae for one-half hour to two hours. Controls were exposed to water used to collect cercariae in an identical manner, and punctured to simulate cercarial tunnels. Fluid from both control and experimental samples was analyzed by LC/MS/MS using a linear ion trap in "triple play" mode. The coexistence of proteins released by cercariae and host skin proteins from epidermis and basement membrane confirmed that cercarial tunnels in skin were sampled. Among the abundant proteins secreted by cercariae was the cercarial protease that has been implicated in degradation of host proteins, secreted proteins proposed to mediate immune invasion by larvae, and proteins implicated in protection of parasites against oxidative stress. Components of the schistosome surface tegument, previously identified with immune serum, were also released. Both lysis and apoptosis of epidermal cells took place during cercarial invasion of the epidermis. Components of lysed epidermal cells, including desmosome proteins which link cells in the stratum granulosum and stratum spinosum, were identified. While macrophage-derived proteins were present, no mast cell or lymphocyte cytokines were identified. There were, however, abundant immunoglobulins, complement factors, and serine protease inhibitors in skin. Control skin samples incubated with water for the same period as experimental samples ensured that invasion-related proteins and host protein fragments were not due to nonspecific degeneration of the skin samples. CONCLUSIONS: This analysis identified secreted proteins from invasive larvae that are released during invasion of human skin. Analysis of specific host proteins in skin invaded by cercariae served to highlight both the histolytic events facilitating cercarial invasion, and the host defenses that attempt to arrest or retard invasion. Proteins abundant in psoriatic skin or UV and heat-stressed skin were not abundant in skin invaded by cercariae, suggesting that results did not reflect general stress in the surgically removed skin specimen. Abundant immunoglobulins, complement factors, and serine protease inhibitors in skin form a biochemical barrier that complements the structural barrier of the epidermis, basement membrane, and dermis. The fragmentation of some of these host proteins suggests that breaching of host defenses by cercariae includes specific degradation of immunoglobulins and complement, and either degradation of, or overwhelming the host protease inhibitor repertoire.