Specificity and kinetics of the milk-clotting enzyme from cardoon (Cynara cardunculus L.) toward bovine .kappa.-casein

The action of Cynara cardunculus L. protease on whole bovine K-casein, over a 3-h period at pH 6.4, was investigated. RpHPLC of the 3% trichloroacetic acid (TCA)-solublefraction of the K-casein digestion mixture showed three peptide peaks, which were identified by amino acid analysis and N-terminal analysis as the 106-169 fragment [caseinomacropeptide (CMP)]. Upon selective precipitation with 12% TCA, one glycosylated and two nonglycosylated forms of CMP were distinguished. Analysis of the whole digestion mixture showed no additional peptides. The kinetics of hydrolysis of the PhelO5- Met106 bond was studied by spectrofluorometry, using fluorescein isothiocyanate-labeled K-casein (FTC- K-casein). The values obtained for kat, k, and k were 1.04 s-l, 0.16 pM, and 6.5 pM-l s-l, respectively. The proteolytic coefficient is of the same order of magnitude as those obtained for other milk-clotting enzymes, but the k, is significantly lower, which reflects the higher affinity of Cynara protease to K-casei

Atividade ovicida e larvicida in vitro do óleo essencial de Eucalyptus globulus sobre Haemonchus contortus.

O objetivo deste trabalho foi avaliar o efeito do óleo essencial de Eucalyptus globulus (OEEG) sobre a eclosão e desenvolvimento de larvas de Haemonchus contortus. A determinação da composição química do OEEG foi feita por meio de cromatografia gasosa acoplada à espectrometria de massa. No teste de eclosão de ovos (TEO), utilizaram-se as concentrações 21,75; 17,4; 8,7; 5,43 e 2,71 mg.mL?1; e no de desenvolvimento larvar (TDL) as concentrações foram 43,5; 21,75; 10,87; 5,43 e 2,71 mg.mL?1. Cada ensaio foi acompanhado por um controle positivo, 0,02 mg.mL?1 tiabendazol, no TEO e 0,008 mg.mL?1 ivermectina no TDL, e por um controle negativo com o diluente Tween 80 (3%). A eficácia máxima obtida pelo OEEG sobre ovos foi de 99,3% na concentração de 21,75 mg.mL?1 e, sobre larvas, 98,7% na concentração de 43,5 mg.mL?1. A concentração do OEEG, que inviabilizou 50% dos ovos e das larvas do parasito, foi de 8,3 e 6,92 mg.mL?1, respectivamente. A análise química do óleo identificou como principal componente o monoterpeno 1,8-cineol. O OEEG apresentou atividade ovicida e larvicida in vitro sobre H. contortus, determinando um bom potencial para utilização no controle de nematóides gastrintestinais de ovinos e caprinos

Atividade ovicida e larvicida in vitro do óleo essencial de Eucalyptus globulus sobre Haemonchus contortus.

O objetivo deste trabalho foi avaliar o efeito do óleo essencial de Eucalyptus globulus (OEEG) sobre a eclosão e desenvolvimento de larvas de Haemonchus contortus. A determinação da composição química do OEEG foi feita por meio de cromatografia gasosa acoplada à espectrometria de massa. No teste de eclosão de ovos (TEO), utilizaram-se as concentrações 21,75; 17,4; 8,7; 5,43 e 2,71 mg.mL-1; e no de desenvolvimento larvar (TDL) as concentrações foram 43,5; 21,75; 10,87; 5,43 e 2,71 mg.mL-1. Cada ensaio foi acompanhado por um controle positivo, 0,02 mg.mL-1 tiabendazol, no TEO e 0,008 mg.mL-1 ivermectina no TDL, e por um controle negativo com o diluente Tween 80 (3%). A eficácia máxima obtida pelo OEEG sobre ovos foi de 99,3% na concentração de 21,75 mg.mL-1 e, sobre larvas, 98,7% na concentração de 43,5 mg.mL-1. A concentração do OEEG, que inviabilizou 50% dos ovos e das larvas do parasito, foi de 8,3 e 6,92 mg.mL-1, respectivamente. A análise química do óleo identificou como principal componente o monoterpeno 1,8-cineol. O OEEG apresentou atividade ovicida e larvicida in vitro sobre H. contortus, determinando um bom potencial para utilização no controle de nematóides gastrintestinais de ovinos e caprinos. Ovicidal and larvicidal activity in vitro of Eucalyptus globulus essential oils on Haemonchus contortus. Abstract: The objective of this work was to evaluate ovicidal and larvicidal effects of Eucalyptus globulus essential oil (EGEO) on Haemonchus conforms. The chemical composition determination of EGEO was through gas chromatography and mass spectrometry. Egg hatch test (EHT) was performed in concentrations 21.75; 17.4; 8.7; 5.43 e 2.71 mg.mL(-1). In larval development test (LDT) were used the concentrations 43.5; 21.75; 10.87; 5.43 e 2.71 mg.mL(-1). Each trial was conducted by negative control with Tween 80 (3%) and positive control, 0.02 mg.mL(-1) of thiabendazole in EHT and 0.008 mg.mL(-1) of ivermectin in LDT. The maximum effectiveness of EGEO on eggs was 99.3% in concentration of 21.75 mg.mL(-1) and on larvae was 98.7% in concentration 43.5 mg.mL(-1). The concentration of EGEO that inhibits 50% of the eggs and larvae was 8.3 and 6.92 mg.mL(-1), respectively. The oil chemical analysis identified as main component the monoterpen 1,8-cineol. EGEO presented ovicidal and larvicidal activities in vitro, revealing a good potential for use in the control of sheep and goat gastrointestinal nematodes

Pervasive gaps in Amazonian ecological research

Biodiversity loss is one of the main challenges of our time,1,2 and attempts to address it require a clear understanding of how ecological communities respond to environmental change across time and space.3,4 While the increasing availability of global databases on ecological communities has advanced our knowledge of biodiversity sensitivity to environmental changes,5,6,7 vast areas of the tropics remain understudied.8,9,10,11 In the American tropics, Amazonia stands out as the world's most diverse rainforest and the primary source of Neotropical biodiversity,12 but it remains among the least known forests in America and is often underrepresented in biodiversity databases.13,14,15 To worsen this situation, human-induced modifications16,17 may eliminate pieces of the Amazon's biodiversity puzzle before we can use them to understand how ecological communities are responding. To increase generalization and applicability of biodiversity knowledge,18,19 it is thus crucial to reduce biases in ecological research, particularly in regions projected to face the most pronounced environmental changes. We integrate ecological community metadata of 7,694 sampling sites for multiple organism groups in a machine learning model framework to map the research probability across the Brazilian Amazonia, while identifying the region's vulnerability to environmental change. 15%–18% of the most neglected areas in ecological research are expected to experience severe climate or land use changes by 2050. This means that unless we take immediate action, we will not be able to establish their current status, much less monitor how it is changing and what is being lost

Caseinolytic Specificity of Cardosin, an Aspartic Protease from the Cardoon Cynara cardunculus L.: Action on Bovine αs- and β-Casein and Comparison with Chymosin

The action of cardosin on bovine αs- and β-casein at 30 °C in 50 mM citrate buffer (pH 6.2) was studied. Peptides were isolated by reversed-phase HPLC on C18 columns and identified from their amino acid composition and N-terminal amino acid sequence. The relative susceptibility of peptide bonds cleaved was Phe23-Phe24 > Trp164-Tyr165 > Tyr166-Val167 > Tyr165-Tyr166 > Phe153-Tyr154 > Phe145-Tyr146 ≈ Leu149-Phe150 ≈ Leu156-Asp157 ≈ Ala163-Trp164 for αs1-casein and Leu192-Tyr193 > Leu191-Leu192 ≈ Leu165-Ser166 > Phe190-Leu191 ≥ Ala189-Phe190 ≈ Leu127-Thr128 for β-casein. In αs2-casein, cardosin cleaved the bonds Phe88-Tyr89 and Tyr95-Leu96. The enzyme shows a clear preference for bonds between hydrophobic, bulky amino acids, cleaving four consecutive peptide bonds in extremely bulky, hydrophobic regions of both αs1-CN (Ala163-Val167) and β-CN (Ala189-Tyr193), which was less attacked by chymosin in various experimental conditions. The active site cleft of cardosin accommodates sequences as bulky as Trp-Tyr-Tyr in different subsites (S1 to S‘2, S2 to S‘1, and probably S3 to S1). Several bitter peptides were identified in the digests