In the present study we investigate the biochemical properties of the members of NPP family in synaptosomes prepared from rat heart left ventricles. Using p-nitrophenyl-5'-thymidine monophosphate (p-Nph-5'-TMP) as substrate for E-NPPs in rat cardiac synaptosomes, we observed an alkaline pH dependence, divalent cation dependence and the K ( M ) value corresponded to 91.42 +/- 13.97 microM and the maximal velocity (V ( max )) value calculated was 63.79 +/- 3.59 nmol p-nitrophenol released/min/mg of protein (mean +/- SD, n = 4). Levamisole (1 mM), was ineffective as inhibitor of p-Nph-5'-TMP hydrolysis in pH 8.9 (optimum pH for the enzyme characterized). Suramin (0.25 mM) strongly reduced the hydrolysis of p-Nph-5'-TMP by about 46%. Sodium azide (10 and 20 mM) and gadolinium chloride (0.3 and 0.5 mM), E-NTPases inhibitors, had no effects on p-Nph-5'-TMP hydrolysis. RT-PCR analysis of left ventricle demonstrated the expression of NPP2 and NPP3 enzymes, but excluded the presence of NPP1 member. By quantitative real-time PCR we identified the NPP3 as the enzyme with the highest expression in rat left ventricle. The demonstration of the presence of the E-NPP family in cardiac system, suggest that these enzymes could contribute with the fine-tuning control of the nucleotide levels at the nerve terminal endings of left ventricles that are involved in several cardiac pathologies.

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The leaves of Quillaja brasiliensis, a native tree species of southern Brazil, accumulate saponins which have adjuvant activity in vaccines. An efficient micropropagation system for Q. brasiliensis was developed. Plants were obtained from aseptically germinated seeds. High rates of germination were observed on filter paper and aseptic medium substrate (sucrose and light exposure did not affect the germination response). Stem nodal segments were inoculated into Murashige and Skoog (MS) medium supplemented with 6-benzylaminopurine. Each explant produced multiple shoots, which elongated satisfactorily on the same medium. Elongated shoots were rooted in a modified MS medium with or without auxins. Best rooting responses were observed in medium containing 10 mg l −1 indole-3-acetic acid under continuous exposure. The rooted explants were acclimatized and successfully transferred to soil, yielding approximately 95% survival after 10 mo. Leaf content of immunoadjuvant saponins in micropropagated plants was not affected by auxin type used for rooting and was comparable to that of field-grown trees. The results indicate that Q. brasiliensis represents an alternative and readily renewable source of biomass for the production of bioactive saponins.