Implantation and Recovery of Long-Term Archival Transceivers in a Migratory Shark with High Site Fidelity

We developed a long-term tagging method that can be used to understand species assemblages and social groupings associated with large marine fishes such as the Sand Tiger shark Carcharias taurus. We deployed internally implanted archival VEMCO Mobile Transceivers (VMTs; VEMCO Ltd. Nova Scotia, Canada) in 20 adult Sand Tigers, of which two tags were successfully recovered (10%). The recovered VMTs recorded 29,646 and 44,210 detections of telemetered animals respectively. To our knowledge, this is the first study to demonstrate a method for long-term (~ 1 year) archival acoustic transceiver tag implantation, retention, and recovery in a highly migratory marine fish. Results show low presumed mortality (n = 1, 5%), high VMT retention, and that non-lethal recovery after almost a year at liberty can be achieved for archival acoustic transceivers. This method can be applied to study the social interactions and behavioral ecology of large marine fishes.     

Isolation of isoflavones from soy-based fermentations of the erythromycin-producing bacterium Saccharopolyspora erythraea

A search for an abundant and economical source of isoflavones, particularly genistein, led to the discovery that the erythromycin-producing organism Saccharopolyspora erythraea also produces this promising new cancer-prevention agent. Erythromycin fermentation is a large-scale, soybean-based process used world-wide for the commercial production of this medically important antibiotic. Results from this study indicate that genistin (the glucoside form of genistein), which is added to the fermentation in the soybean media, was converted to genistein through the action of a β-glucosidase produced by the organism. Genistein was co-extracted with erythromycin from the fermentation broth, then separated from erythromycin during the second step of the purification process for the production of erythromycin. Received 10 September 1996 / Received revision: 22 November 1996 / Accepted: 7 December 1996     

Determination of small quantities of sulfate (0-12 nmol) in serum, urine, and cartilage of the mouse

The colorimetric benzidine method of K. S. Dodgson and B. Spencer (1953, Biochem. J. 55, 436-440) for the measurement of inorganic sulfate can be scaled down about 100 times by using disposable 96-well microplates instead of individual cuvettes. Ten-microliter samples of serum and urine, derived from mice, can be analyzed in a simple, rapid, and reliable way without sacrificing the animals. Without prior isolation of sulfated glycosaminoglycans, ester sulfate in mouse patellar cartilage is liberated quantitatively as inorganic sulfate upon acid hydrolysis in 3 M HCl for 16 h at 80 degrees C. To this end the articular cartilage layer of the patella must be separated in toto from the underlying bone. Subsequent hydrolysis in polypropylene tubes gives accurate results. In contrast, hydrolysis in borosilicate glass vials is useless, since nanomoles of sulfate added cannot be recovered adequately. The thin patellar cartilage layer obtained from 10-week-old male mice contains about 5 nmol of sulfate, an amount easily measured with the developed microplate benzidine method.     

Low nanogram detection of nucleotides using fast atom bombardment-mass spectrometry

The effect of trimethylsilyl (TMS) derivatization on detection limits of mononucleotides in fast atom bombardment-mass spectrometry (FAB-MS) was examined. FAB-MS methods were developed to optimize sensitivity using adenosine 5'-monophosphate as a model compound and then applied to reference standards of two clinically important nucleotides: tricyclic nucleoside-5'-monophosphate (TCNMP) and 5-fluoro-2'-deoxyuridine-5'-monophosphate (FdUMP). The detection limit for the TMS derivative of TCNMP was 2.5-5 ng/microliters and less than 2.5 ng/microliters for FdUMP as its TMS derivative. This is greater than two orders of magnitude more sensitive than the FAB-MS analysis of the corresponding free compounds. These low detection limits for the TMS derivatives were obtained using a narrow scan range, signal averaging, detection in the negative ion mode, and 3-nitrobenzyl alcohol as the matrix. Hydrolysis of one or more of the labile TMS groups did occur, with the extent of hydrolysis being greatest in the more protic matrices.     

Circadian Variations in the Affinities of Nad Kinase and Nadp Phosphatase for their Substrates, Nad+ and Nadp+ in Dividing and Nondividing Cells of the Achlorophyllous Zc Mutant of Euglena Gracilis Klebs (Strain Z)

-We have previously shown that NAD kinase and NADP phosphatase activities display circadian rhythms, in the soluble (SN) and membrane-bound (P) fractions of crude extracts of the achlorophyllous ZC mutant of the phytoflagellate Euglena gracilis (which displays circadian rhythmicity of cell division). We determined if changes in the affinity of NADP phosphatase and NAD kinase for their substrates, NADP⁺ and NAD⁺, were occurring by calculating the ratios 100(velocity found in Km conditions/velocity found in saturating conditions). The rationale was that if the affinity remained unchanged according to circadian time (CI), these values should always equal 50, independently of any changes in enzyme quantity; values greater than 50 should indicate increases in enzyme affinity, and values less than 50 decreases in affinity. Our results indicated that these values calculated for NADP phosphatase exhibited a complex pattern of rhythmicity, while those for NAD kinase displayed circadian variations strongly correlated with the rhythms in enzyme activity. The curves showed troughs at CT 00-04 both in dividing and nondividing cells and peaks at CT 18-20 or at CT 08-14 in cells sampled, respectively, from a dividing or a stationary culture. Such variations are indicative of changes in the kinetic properties of the enzyme, which may reflect modifications in its affinity either for effectors (such as Ca2⁺-calmodulin) or for its substrate, NAD⁺. This may be due to (i) the expression of different isoenzymes at different CTs; (ii) different posttranslational modifications of the enzyme; or (iii) concentrations of effectors varying in a circadian manner.