Although transcription and transcript processing in the chloroplasts of plants have been extensively characterised, the RNA metabolism of other chloroplast lineages across the eukaryotes remains poorly understood. In this paper, we use RT-PCR to study transcription and transcript processing in the chloroplasts of Amphidinium carterae, a model peridinin-containing dinoflagellate. These organisms have a highly unusual chloroplast genome, with genes located on multiple small 'minicircle' elements, and a number of idiosyncratic features of RNA metabolism including transcription via a rolling circle mechanism, and 3' terminal polyuridylylation of transcripts. We demonstrate that transcription occurs in A. carterae via a rolling circle mechanism, as previously shown in the dinoflagellate Heterocapsa, and present evidence for the production of both polycistronic and monocistronic transcripts from A. carterae minicircles, including several regions containing ORFs previously not known to be expressed. We demonstrate the presence of both polyuridylylated and non-polyuridylylated transcripts in A. carterae, and show that polycistronic transcripts can be terminally polyuridylylated. We present a model for RNA metabolism in dinoflagellate chloroplasts where long polycistronic precursors are processed to form mature transcripts. Terminal polyuridylylation may mark transcripts with the correct 3' end.

Subjects were each given either a 25, 50 or 100 mg intravenous loading dose of oxpentifylline followed by an intravenous infusion at a constant rate of 1.5 mg/min for 3 h. Plasma levels of oxpentifylline were measured to obtain information on its pharmacokinetics and to establish which of the loading doses gave the most rapid attainment of the steady state plasma levels of intact drug. Oxpentifylline kinetics were best described by a two compartment model giving a characteristic dip in the plasma level versus time curves before steady state was reached when either the 50 or 100 mg loading doses, followed by the constant intravenous infusion, were given. The terminal half-life of oxpentifylline was 1.02 +/- 0.86 h, reflecting a very high clearance of the drug (approx. 3 000 to 6 000 ml/min). The high clearance could be attributed to extrahepatic metabolism occurring in blood which was observed in vitro using whole blood but not plasma. The clearance of a reduced metabolite of oxpentifylline was less than that of the intact drug, although the half-life was similar (0.83 +/- 0.18 h). Of the three loading doses tested, only the highest showed any side effects, these being transient and occurring within a 5 to 10 min period after dosing and appeared to correlate with the high initial plasma levels of the drug. The 25 mg loading dose gave initial plasma levels generally below the final steady state levels, whilst the 50 mg loading was the closest to giving immediate steady state plasma levels of oxpentifylline.

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Paraffin wax embedding methods suitable for demonstrating the distribution of enzyme activity in tissues sections are uncommon; most procedures rely on the use of frozen section techniques. This paper describes a system for demonstrating certain enzymes which involves incubation of the tissue with appropriate substrates before a Paramat wax embedding procedure. While it has distinct merits of its own, the procedure is eminently suitable for use where a cryostat is not available; it can also be readily applied to other enzymes and tissues.

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