Active site mapping of Loxosceles phospholipases D: Biochemical and biological features.

Brown spider phospholipases D from Loxosceles venoms are among the most widely studied toxins since they induce dermonecrosis, triggering inflammatory responses, increase vascular permeability, cause hemolysis, and renal failure. The catalytic (H12 and H47) and metal-ion binding (E32 and D34) residues in Loxosceles intermedia phospholipase D (LiRecDT1) were mutated to understand their roles in the observed activities. All mutants were identified using whole venom serum antibodies and a specific antibody to wild-type LiRecDT1, they were also analyzed by circular dichroism (CD) and differential scanning calorimetry (DSC). The phospholipase D activities of H12A, H47A, H12A-H47A, E32, D34 and E32A-D34A, such as vascular permeability, dermonecrosis, and hemolytic effects were inhibited. The mutant Y228A was equally detrimental to biochemical and biological effects of phospholipase D, suggesting an essential role of this residue in substrate recognition and binding. On the other hand, the mutant C53A-C201A reduced the enzyme's ability to hydrolyze phospholipids and promote dermonecrosis, hemolytic, and vascular effects. These results provide the basis understanding the importance of specific residues in the observed activities and contribute to the design of synthetic and specific inhibitors for Brown spider venom phospholipases D.

Structural Insights into Substrate Binding of Brown Spider Venom Class II Phospholipases D.

Phospholipases D (PLDs), the major dermonecrotic factors from brown spider venoms, trigger a range of biological reactions both in vitro and in vivo. Despite their clinical relevance in loxoscelism, structural data is restricted to the apo-form of these enzymes, which has been instrumental in understanding the functional differences between the class I and II spider PLDs. The crystal structures of the native class II PLD from Loxosceles intermedia complexed with myo-inositol 1-phosphate and the inactive mutant H12A complexed with fatty acids indicate the existence of a strong ligand-dependent conformation change of the highly conserved aromatic residues, Tyr 223 and Trp225 indicating their roles in substrate binding. These results provided insights into the structural determinants for substrate recognition and binding by class II PLDs.

[Chronic hepatitis C. Epidemiology and therapeutic results in 255 cases]. / Hepatitis crónica por virus C. Datos epidemiológicos y de eficacia terapéutica en 255 casos.

A review of the statistics of the years 2003, 2004 to August 2005 has provided data corresponding to 255 patients that were diagnosed and included in the treatment protocol of chronic hepatitis C. Epidemiologically, the 2/1 man/woman ratio, mean age of 43 years and history of transfusion (transf.) in 18.6% are to be noted. Parenteral drug addicition (PDA) in 17% and alcoholism in 14.1%, of unknown origin in 58.74% of the cases. 79.05% corresponded to genotype 1, 4.74% to genotype 4, 14.22% to genotype 3, and 2.76% to genotype 2. Genotypes 1 and 4 were included in the treatment regime with PEG-Interferon alpha-2b + Ribavirin (with doses adjusted for weight) and those of genotype 3 and 2 in the regime with Interferon alpha 2b + Ribavirin. In 95 cases of Genotype 1 (87) and 4 (8), a sustained viral response (SVR) of 61.95% was obtained, with three drop-outs. In the 20 cases of genotypes 3 (17) and 2 (3), SVR was 85%, with no drop-outs. In the 20 cases of genotypes 3 (17) and 2 (3), SVR was 85%, with no drop-outs. The adjustment of treatment to body mass involved an economic savings per case amounting to 868.94 Euro. Finally, in 16 cases of Genotype 1, before treated with Interferon in monotherapy, a sustained viral response of 31.25% was obtained with combined therapy.

Quantification of astaxanthin in shrimp waste hydrolysate by HPLC.

In the present study, a simple and rapid reversed-phase HPLC method for the determination of astaxanthin in shrimp waste hydrolysate has been developed and validated. The analytical procedure involves the direct extraction of astaxanthin from the lipid fraction with methanol. The analytical column, SS Exil ODS, was operated at 25C. The mobile phase consisted of a mixture of water:methanol:dichloromethane:acetonitrile (4.5:28:22:45.5 v/v/v/v) at a flow rate of 1.0 mL/min. Detection and identification were performed using a photodiode array detector (lambda(detection) = 476 nm). The proposed HPLC method showed adequate linearity, repeatability and accuracy.

[Latent sprue in a 62-year-old man]. / Esprue latente en varón de 62 años.

We report a case of a 62-years-old male, diagnosed by radiology, antigliadin antibodies and intestinal biopsy of sprue, without fat malabsorption. The patient with a free gluten diet evolved to a clinic and radiologic normalization in three months and a great histological improvement.