Quinine and quinidine production by cinchona leaf,root and unorganized cultures

Serially propagated Cinchona ledgeriana and C. succirubra (Rubiaceae) leaf, root and unorganized suspension cultures established from germinated seeds were studied for quinine and quinidine production. Leaf organ cultures were grown and subcultured in Murashige and Skoog's Revised Tobacco Medium supplemented with benzyladenine; root organ cultures were grown on the same medium supplemented with indolebutyric acid; and unorganized suspension cultures were grown on the same medium supplemented with 2,4-dichlorophenoxyacetic acid and benzyladenine. On a dry weight basis, leaf organ cultures of C. ledgeriana contained 0.06 % quinine and 0.05 % quinidine and of C. succirubra contained 0.04 % quinine and 0.04 % quinidine. No quinine and quinidine were detected in either root organ or unorganized suspension cultures.     

Response of Guanosine 5′-Triphosphate Concentration to Nutritional Changes and Its Significance for Bacillus subtilis Sporulation

We have investigated the changes in the guanosine 5'-triphosphate (GTP) and P-ribosyl-PP pools in stringent and relaxed strains of Bacillus subtilis under conditions frequently used to initiate sporulation. After a shift-down from a Casamino Acids-glutamate to a glutamate medium (Sterlini-Mandelstam shift-down), the pools of adenosine 5'-triphosphate and P-ribosyl-PP increased in both strains; in the stringent strain, ppGpp and pppGpp increased and GTP decreased rapidly, whereas in the relaxed strain, ppGpp and pppGpp increased only slightly and GTP decreased only slowly and less extensively. The stringent strain sporulated well, whereas the relaxed strain sporulated late and poorly. Addition of decoyinine, an inhibitor of guanosine 5'-monophosphate synthetase, caused a further decrease of GTP and initiated good sporulation of the relaxed strain. After a shift-down from a glucose-lactate to a lactate medium (Ramaley-Burden shift-down) the pool of P-ribosyl-PP (and GTP) decreased in both strains, indicating a shortage of purine precursors. This shift-down also caused a stringent response which prevented the consumption of nucleotides, as shown by the maintenance of adenosine 5'-triphosphate at a high concentration in the stringent strain but not in the relaxed strain. After a delay, the relaxed strain, in which GTP decreased as fast as in the stringent strain, sporulated also as efficiently. In nutrient sporulation medium the stringent strain and, less effectively, the relaxed strain accumulated ppGpp and pppGpp transiently towards the end of exponential growth. Eventually, the P-ribosyl-PP pool decreased drastically in both strains. In all cases the initiation of sporulation was correlated with a significant decrease of GTP. Granaticin, an antibiotic which prevents the charging of leucyl-transfer ribonucleic acid, was used to show that the stringent response inhibited the formation of xanthosine monophosphate from inosine monophosphate. It prevented the accumulation of xanthosine monophosphate in decoyinine-treated cultures of the stringent strain but not in those of the relaxed strain.     

Evaluation of Bone Strength During Aflatoxicosis and Ochratoxicosis

Young chickens were fed graded levels of aflatoxin (0, 0.625, 1.25, 2.5, 5.0, and 10.0 μg/g of diet) or ochratoxin (0, 0.5, 1.0, 2.0, 4.0, and 8.0 μg/g of diet), and the breaking strength, displacement before failure, and diameter of their tibias were determined. Breaking strength was decreased at growth inhibitory levels of aflatoxin (2.5 μg/g) and ochratoxin (2 μg/g), whereas a reduction in diameter required higher levels (5.0 and 4.0 μg/g, respectively). Bones from birds with ochratoxicosis selected to have diameters equal to control bones had lower breaking strength. In an attempt to negate mathematically the effect of decreased diameter and bias in any selection process, stress at time of failure of the bones was calculated and found to be decreased by feeding aflatoxin but not ochratoxin. Total displacement of bones before breaking was increased significantly (P < 0.05) by both toxins at the highest levels administered, but this increase was primarily the result of an increase in displacement from the start of failure to complete failure. Increased displacement associated with both toxicoses was equal in bones selected to be of equal diameter or in bones from the same treatment but of different diameters. However, calculation of modulus of elasticity which is corrected for diameter revealed aflatoxin had no effect whereas ochratoxin tripled the effect. These data indicate that the material properties of bones can be altered during mycotoxicoses and suggest yet another way in which mycotoxins are detrimental to animal health.     

TRANSMITTER METABOLIZING ENZYMES AND FREE AMINO ACID LEVELS IN SENSORY AND MOTOR NERVES AND GANGLIA OF THE SHORE CRAB (CARCINUS MAENAS)

—The distribution of ChAT (choline acetyltransferase), GAD (glutamate decarboxylase) and acetylcholinesterase in some sensory and motor nerves of the shore crab, Carcinus maenas, has been investigated using micro-assay techniques. ChAT was concentrated in the afferent nerve fibres of the thoracic-coxal muscle receptor as well as in the coxo-basal chordotonal receptor nerve and other leg sensory fibres. GAD was found in leg motor nerves including the promotor and remotor muscle nerves, being undetectable in the sensory nerves. Acetylcholinesterase was found in similar levels in both sensory and motor nerves assayed. Amino acid analysis using a micro-dansylation technique showed that sensory nerves had low GABA levels, whereas the leg nerve including motor fibres had substantially higher GABA concentrations. GAD and GABA were also found in low amounts in the leg promoter mucle, which is consistent with GABA being a neuromuscular transmitter.     

Cooperative control of translational fidelity by ribosomal proteins in Escherichia coli

Summary Two spontaneous mutants of Escherichia coli strain KMBL-146 selected for resistance to the aminoglycoside antibiotic neamine show severe restriction of amber suppressors in vivo. Purified ribosomes from the mutant strains exhibit low neamine-induced misreading in vitro and a decreased affinity for the related antibiotic streptomycin.Biochemical analysis shows that the mutants each have two modified 30S ribosomal proteins, S12 and S5. In agreement with these results, genetic analysis shows that two mutations are present, neither of which confers resistance to neamine by itself; the mutation located in gene rpxL (the structural gene for protein S12) confers streptomycin dependence but this dependence is suppressed in the presence of the second mutation, located in gene rpxE (the structural gene for protein S5).     

Acetyl groups in cell-wall preparations from higher plants

O-Acetyl groups were detected by i.r. spectroscopy in cell-wall preparations from grasses and other higher plants and their presence was confirmed chemically. The amounts present are likely to influence both the physical state of the cell-wall polysaccharides and also their digestion by enzymes.     

Tritium labeling of thermolysin, elastase, and ribonuclease by exposure to tritium gas at low pressure

The bacterial metalloendoprotease thermolysin, bovine pancreatic ribonuclease, and porcine pancreatic elastase have been tritiated by exposure to subcurie amounts of tritium gas at pressures below 50 mTorr for periods of 1 to 6 h. Thermolysin, ribonuclease, and elastase have been purified to specific radioactivities of 15, 5, and 1 Ci/mol, respectively. Amino acid analyses of the tritiated enzymes revealed higher relative specific radioactivities for His, Pro, and Phe in all three proteins while Val and Ile were among the residues with the lowest relative specific radioactivities. The recovery of enzyme activity was always greater than 95% and the formation of tritiated decomposition products was not observed. This lowpressure gas exposure process requires less tritium gas and less time than the original method of Wilzbach to achieve equal or higher levels of tritium incorporation. In addition, the enzymes were completely active and did not show the presence of highly radioactive byproducts which have been observed in earlier studies of the Wilzbach labeling of proteins.