(1): equation(1) CM=∑IλFλ/∑Fλ,CM=∑IλFλ/∑Fλ,where
Fλ stands for the fluorescence emission at wavelength Iλ and the summation was carried out over the range of appreciable values of F. Far-UV CD spectra were recorded in the 190–250 nm region, in a 1-mm path length quartz cuvette using a spectropolarimeter (JASCO J-810). The instrument was calibrated with D-10-camphorsulfonic acid. The protein concentrations were as follows: non-irradiated (8 μM), 1 (8 μM), 10 (10 μM) and 25 kGy (20 μM) for WGA, in phosphate buffer, pH 7.2 at 25 °C. After irradiation the samples were centrifuged and the measurements were performed this website with the supernatant. The data were averaged for eight scans that were performed at a speed of 50 nm/min and collected in 0.5-nm steps. The baselines (buffer alone) were subtracted from the protein spectra. Results were expressed as mean residue ellipticity, [θ], defined as [θ]=θobs/(10.C.l.n.),[θ]=θobs/(10.C.l.n.),where θobs is the CD in millidegrees, C is the protein concentration (M), l is the path-length of the cuvette (cm) and n is the number of amino acid residues assuming a mean number of 186 residues. Female Swiss albino mice (5 weeks old) were obtained from the breeding colony of the Departamento de Antibióticos da Universidade Federal de
CDK activation Pernambuco, Brazil, and given ad libitum access to food and water. The animals were kept in an environmentally controlled room, temperature 21 ± 2 °C, under a light/dark cycle of 12 h. Requirements for care and handling of experimental animals were according to international and Brazilian regulations. All test substances were administered intragastrically by tube. WGA was dissolved in 0.5 mL of 0.9% sterile saline. Swiss
albino mice were immunised subcutaneously on Day 0, 15 and 30, using 0.5 ml WGA (10 μg/mL) dissolved in saline without use of an adjuvant (five mice per group). Control animals were treated subcutaneously with 0.5 mL Lepirudin saline. Three days before starting the oral treatment in animals, they were stimulated with the same dose intraperitoneally. Over 7 days, mice were treated thus: group A, immunised mice were treated with 1 mL saline/day; group B, immunised mice were treated with non-irradiated WGA; group C and D immunised mice were treated with irradiated WGA at 1 and 10 kGy, respectively. The dose of WGA (27 mg/kg body weight/day) was according to total dietary intake of lectins in human subjects consuming vegetarian diets (calculations based on data from Peumans & Van Damme, 1996). Body weight was determined before and after immunisation and after oral challenge. The final body weight of each group was obtained from the means of the individual values and expressed in grams. Blood samples were obtained and placed into micro-blood tubes containing the anticoagulant ethylenediaminetetraacetic acid (EDTA).