Multiple phosphorylation of acetyl-CoA carboxylase in chick liver cells. A cyclic AMP-independent process. When chick liver cells in monolayer culture were incubated with 32Pi in the presence of insulin, acetyl-CoA carboxylase became extensively labeled with 32Pi reaching a stoichiometry of 9 to 10 mol of phosphoryl group per mol of 240,000-dalton enzyme subunit. The covalently bound phosphate was found to be metabolically labile, turning over with a t1/2 of approximately 2 h (enzyme t1/2 approximately equal to 24 h). Addition of Bt2cAMP altered neither the rate nor extent of phosphorylation. Contrary to other reports, the fully phosphorylated acetyl-CoA carboxylase appears to be catalytically active.


Metabolism of prostacyclin in blood vessels. The activity of 15-hydroxyprostaglandin dehydrogenase has been shown to be high in both mesenteric arteries and veins; the present study suggests that it may be responsible for the inactivation of prostacyclin (PGI2). The cytoplasmic fractions of bovine mesenteric arteries and veins were incubated with radiolabeled PGI2 in the presence of NAD+ or NADP+. The substrate was rapidly converted to a product, which was isolated and identified as 6,15-diketo prostaglandin F1alpha, (6,15-diketo-PGF1alpha) by thin layer chromatography and gas chromatography-mass spectrometry. The initial reaction rate began to level off after less than 1 min of incubation at 37 degrees C. When radiolabeled 6-keto-PGF1alpha, the stable hydrolysis product of PGI2, was used as substrate under the same conditions, 97% was recovered unmetabolized after 2 min of incubation. Catabolism of PGI2 may be a major determinant of its levels in blood vessels and, therefore, may be of crucial importance to regulating the action of PGI2. Further, estimation of PGI2 generation by either tissues or organs may be misleading if only 6-keto-PGF1alpha is measured.


Effects of pH and temperature on the interaction of an impermeant probe with surface proteins of the human red blood cell. The conformation of the outer surface of the human red cell membrane has been studied under various conditions using the impermeant probe [125I]diazodiiodosulfanilic acid. At least seven polypeptides were labeled by the reagent, including the three extractable glycoproteins separable by the electrophoretic method employed. The Mr = 43,000 protein band was shown to contain two labeled species, one a glycoprotein, in addition to its major constituent, red cell actin. The extent and pattern of labeling were very sensitive to changes in pH and temperature. Total labeling increased with increasing pH and was greater at 4 degrees C than 37 degrees C. Binding of the probe to the Mr = 90,000 polypeptide and the major glycoprotein were relatively increased with increasing pH and temperature while opposite effects were observed for the Mr = 43,000 peptide(s). The pH effects on external membrane labeling were rapidly reversible. Results were similar in cells of different densities, suggesting that the pH and temperature effects were not related to cell age. The data presented emphasize the lability of membrane conformation and reactivity and thus the necessity to consider carefully the conditions of labeling in interpretation of studies using impermeant probes.


Solvent effects on the structure of rabbit Clq, a subcomponent of the first component of complement. The effect of solvent conditions on the conformation of rabbit Clq was studied by both spectroscopic and nonspectroscopic methods. The conformation of Clq in buffered saline solutions at pH 7.4 or 6.0 did not differ significantly from Clq at twice the saline concentration as determined with circular dichroism, difference spectroscopy, and tritium-hydrogen exchange techniques. Addition of calcium to the buffers had no structural effects in any of the conditions examined. Hydrogen exchange experiments performed at pH 7.4 were also unaffected by magnesium, manganese, or ethylenediaminetetraacetic acid. With all the methods used a pH effect was observable between 5.1 and 8.3. From solvent perturbation difference spectroscopy results it was calculated that the equivalent of 10 +/- 2 and 6 +/- 1 mol of tyrosine and tryptophan/mol of Clq, respectively, became exposed at the lower pH. A small positive CD band in the 231 to 235 nm region decreased in wavelength and increased in magnitude as a function of decreasing pH, indicating tyrosine exposure at the lower pH and possibly changes in the collagen-like structure of Clq. Hydrogen exchange experiments indicate a small, but significant, conformation transition occurring in the pH 5 region and a stabilization of conformation between pH 6 to 8. From these results the conformational pH dependence was interpreted as an acid expansion of Clq with a minor conformational transition occurring between pH 5 AND 6. These effects may in part be associated with decreased Clq-Ig interactions which have been observed at the lower pH.


Leghemoglobin. Low temperature optical spectra of acid and alkaline forms of leghemoglobin(IV). Configuration of the heme. Leghemoglobin(IV), the derivative of leghemoglobin at the formal oxidation state IV, when cooled to liquid nitrogen temperature exhibits radically different spectra at acid and alkaline pH. The acid and alkaline forms are freely interconvertible. The optical spectrum of the acid form is closely similar to optical spectra of the red higher oxidation states of horseradish and cytochrome c peroxidases, showing that the configuration of the heme iron is the same throughout this family of compounds. That configuration is believed to be Fe(IV) in a porphyrin environment. The optical spectrum of the alkaline form of leghemoglobin(IV) recalls that of alkaline low spin ferric leghemoglobin. Near infrared spectra of leghemoglobin(IV), myoglobin(IV), and the higher oxidation states of the peroxidases are featureless to 1300 nm, suggesting a common structural feature. The acid form of leghemoglobin(IV), seen in fluid buffer as a transient species at pH 5 or less, is conveniently generated by cooling a solution of the more stable alkaline form in borate buffer to liquid nitrogen temperature. At this temperature borate buffers become acid.


Acid and alkaline forms of the higher oxidation state of kangaroo, horse, and sperm whale myoglobin. Myoglobin(IV), the derivative of myoglobin at the formal oxidation state IV, prepared from kangaroo (Megaleia rufa), horse, or sperm whale myoglobin, when cooled to liquid nitrogen temperature, assumes acid and alkaline forms with different optical spectra. The essential features of the optical spectra of the acid forms are the same as those of leghemoglobin(IV) and are very similar to those of optical spectra of the red higher oxidation states of catalases and peroxidases. This shows that the configuration of the heme iron is the same throughout these compounds. That configuration is believed to be Fe(IV) in a porphyrin environment. The optical spectra of alkaline mammalian myoglobin(IV), like that of alkaline leghemoglobin(IV), resemble those of the alkaline low spin ferric proteins. Kangaroo myoglobin(IV) may be prepared by reaction of ferrous myoglobin with hydrogen peroxide. The acid forms of myoglobin(IV) are conveniently prepared by cooling solutions in borate buffers, initially pH 8.3, to liquid nitrogen temperature. At this temperature borate buffers become acidic.


Molecular properties of a major cell surface protein from chick embryo fibroblasts. The molecular structure of chick embryo fibroblast cell surface protein has been investigated by ultracentrifugation, circular dichroism, and fluorescence. Most measurements were restricted to alkaline solutions because of the limited solubility of this protein at more neutral pH values. A very high frictional ratio for the protein suggests an asymmetric structure. However, there are elements of organized structure since typical thermal transition curves were found by several methods. Consequently, a model in which ordered domains are connected by flexible polypeptide chains seems to account for all the hydrodynamic and optical data.


Rupture of the plantar fascia in athletes. Symptoms resembling those of plantar fasciitis were seen in six athletes who were thought to have a partial rupture of the plantar fascia. Treatment, which included the use of crutches, anti-inflammatory agents, strapping of the arch, and ice packs, was successful in all but one patient who had a painful mass in the area of the previous rupture. After surgical excision of the painful mass and release of the fascia, he recovered. Five of the six athletes had been previously treated with repeated local injections of steroid.


Glucocorticoid induction of tyrosine hydroxylase in a continous cell line of rat pheochromocytoma. We have established a continous cell line (G1) in which the tyrosine hydroxylase specific activity is increased as much as 50-100-fold in response to dexamethasone. This response is specific for the glucocorticoid class of steroid hormones; it is elicited by dexamethasone, corticosterone, and triamcinolone, but not by estradiol, testosterone, progesterone, or deoxycorticosterone acetate. The increase in tyrosine hydroxylase specific activity is likely to be due to the increased synthesis of new enzyme protein rather than an activation of existing protein molecules, inasmuch as this increase is completely blocked by cycloheximide.


Inhibition of glucose transport in human erythrocytes by benzylalcohol. The inhibition of glucose transport by the non-ionizable local-anesthetic denzylalcohol is of the mixed type and independent of pH. The affinity of benzylalcohol to the free carrier is about three times larger than that to the carrier-glucose complex.


Culture pH, CO2 tension, and cell division in Euglena gracilis Z. Growth characteristics of Euglena gracilis Z as functions of culture pH, CO2 tension, temperature, and lighting regime were investigated. The results are consistent with the possibility that cell division is preceded by a lowered intracellular pH. Also consistent with this possibility is the finding that division rhythmicity can be induced by periodic changes in CO2 tension. It is suggested that the rhythmicity is induced by changes in intracellular pH produced by carbonic acid.


Effects of acidified fetal bovine serum on the fibrinolytic activity and growth of cells in culture. The fibrinolytic activity of cells in culture varied with the type of serum employed in the growth medium. Degradation of iodinated fibrin occurred slowly when Rous sarcoma virus-transformed chick embryo fibroblasts were grown in medium containing fetal bovine serum (FBS), and rapidly when chicken serum was employed. This difference reflected the low plasminogen and high inhibitor content of FBS. The inhibitors were found to be serum macromolecules that were precipitated with ammonium sulfate or polyethylene glycol, and were inactivated by boiling or upon exposure to acidic conditions. No inhibitor activity was detected in fetuin, one of the major proteins present in FBS. Acidified FBS was similar to chicken serum in that both supported high rates of cell-mediated fibrinolytic activity. Although virally transformed hamster, mouse and chicken cells grew well in acid-treated FBS, their normal counterparts did not. Apparently, acifification resulted in the formation of materials that were toxic to normal cells. These agents rapidly blocked cellular DNA synthesis.


Effects of methylprednisolone on hydrogen ion absorption in the canine stomach. The effect of methylprednisolone (2 mg/kg per day given parenterally for 3 doses, 2 wk or 12 wk) on the permeability of mammalian gastric mucosa to hydrogen ion (H(+)) was examined with denervated fundic pouches in dogs with antrectomies. Transmucosal electric potential difference (PD) and net fluxes of H(+) and Na(+) were determined for luminal [H(+)] from 20 to 160 mM and [Na(+)] from 1 to 140 mM ([H(+)] and [Na(+)] were varied reciprocally). The PD was 50-60 mV lumen negative and was constant over the entire range of Na(+) and H(+) concentration tested. Net H(+) flux varied linearly with [H(+)]. Extrapolation indicated apparent H(+) loss at zero luminal concentration, suggesting a basal HCO(3) (-) secretion. Addition of acetylsalicylic acid (ASA) or taurocholate decreased the PD to 30-40 mV and increased threefold the slope of the relation between net H(+) flux and [H(+)] (k(H)). Calculation of PD-independent permeability constants for H(+) (P(H)) with the Goldman constant field equation indicated that this increase in k(H) could not be attributed solely to the associated decrease in PD. Prednisolone administered for 3 doses had no effect on either the basal mucosal permeability to H(+) or the altered permeability induced by ASA or taurocholate. Chronic administration induced a low rate of basal acid secretion (at 12 wk) but had no effect on either PD or k(H). However, the increase in k(H) and P(H) that developed upon addition of ASA or taurocholate in chronically treated dogs was more than one and a half times that of controls. These data suggest that prolonged treatment with glucocorticoids increases susceptibility of the gastric mucosa to damage by agents that increase permeability to H(+).


The role of acetaldehyde in mediating the deleterious effect of ethanol on pyridoxal 5'-phosphate metabolism. Previous studies in vivo and with isolated perfused rat livers have suggested that the deleterious effect of ethanol on hepatic pyridoxal 5'-phosphate metabolism is mediated by acetaldehyde. Inasmuch as acetaldehyde has no effect on the synthesis of pyridoxal phosphate, it has also been postulated that acetaldehyde accelerates pyridoxal phosphate degradation by displacing this coenzyme from binding proteins, which protect it against hydrolysis. To test these hypotheses, studies have been performed with isolated rat hepatocytes, subcellular fractions of rat liver, and human erythrocytes. Ethanol oxidation lowered the pyridoxal phosphate content of isolated liver cells when acetaldehyde oxidation was inhibited by either disulfiram or prior treatment of rats with cyanamide. Additions of 7.5 mM acetaldehyde alone at 40-min intervals to cell suspensions decreased hepatic pyridoxal phosphate content only slightly because acetaldehyde was rapidly metabolized. However, when acetaldehyde oxidation and reduction were inhibited by cyanamide treatment and by 4-methyl-pyrazole and isobutyramide, respectively, a 40% decrease in hepatic pyridoxal phosphate content was observed in 80 min of incubation. In equilibrium dialysis experiments, acetaldehyde, 7.5 and 15 mM, displaced protein-bound pyridoxal phosphate in undialyzed hepatic cytosol and in hemolysate supernate containing added pyridoxal phosphate. In the presence of alkaline phosphatase, acetaldehyde accelerated the degradation of pyridoxal phosphate in dialyzed hemolysate supernate and hepatic cytosol with added pyridoxal phosphate. Acetaldehyde also inhibits tyrosine aminotransferase. The kinetics of inhibition were mixed competitive-noncompetitive with respect to pyridoxal phosphate. These observations support the hypothesis that the deleterious effect of ethanol oxidation on pyridoxal phosphate metabolism is mediated at least in part by acetaldehyde which displaces this coenzyme from protein binding, thereby enhancing its degradation.


Human erythrocyte hexokinase deficiency. Characterization of a mutant enzyme with abnormal regulatory properties. In the erythrocytes of a patient with hereditary nonspherocytic hemolytic anemia, a homozygous expression of hexokinase deficiency was detected. The mutant enzyme was characterized by normal kinetic parameters with respect to its substrates, glucose and MgATP2-, normal pH optimum, normal heat stability at 40 degrees C, but abnormal behavior with respect to its regulation by glucose-1,6-diphosphate and inorganic phosphate, and an altered electrophoretic pattern. Interpretation of the results revealed the presence of two different hexokinases type I in normal human erythrocytes: one enzyme with a high affinity for glucose-1,6-diphosphate, the inhibition of which is regulated by inorganic phosphate; and another enzyme with a lower affinity for the inhibitor, not regulated by inorganic phosphate. The former enzyme was not detectable in the erythrocytes of the patient, whereas the presence of the latter enzyme could be demonstrated.


Passive transfer by cells of type II collagen-induced arthritis in rats. To investigate the role of immunologic hypersensitivity to collagen in the causation of type II collagen-induced arthritis in rats, passive transfer experiments were performed. Wistar/Lewis rats used in these experiments were demonstrated to be histocompatible by prolonged skin graft survival and mixed lymphocyte cultures. Popliteal lymph node weight assays excluded a potential for graft-vs.-host reactivity in this strain. 9 of 32 naive rats developed arthritis after intravenous receipt of pooled spleen and lymph node cells from donors that had been injected intradermally with type II collagen emulsified in incomplete Freund's adjuvant. This passively transferred synovitis was evident clinically as well as histologically. In control cell transfer experiments involving a total of 97 recipients, transfer of arthritis was shown to require viable cells sensitized to type II collagen. These controls included 17 rats receiving cells from unimmunized donors, 20 recipients of cells from donors injected with incomplete Freund's adjuvant alone, and 24 recipients of cells from rats injected with type I collagen in adjuvant. Deliberate addition of solubilized type II collagen to unsensitized cells at the time of transfer or injection of heat-killed sensitized cells also did not cause arthritis in a total of 36 recipients. These latter two control groups indicate that disease transfer was not the result of antigen carry-over. Intravenous injection of sera from arthritic donors was incapable of passively transferring clinical or histologic synovitis in 30 recipients. Thus, these studies directly implicate immunologic sensitivity to the cartilage type of collagen in the etiology of this autoimmune disease.


The effect of complement depletion on lung clearance of bacteria. We have investigated the effect of hypocomplementemia on early pulmonary clearance of four species of bacteria. The experiments were performed in an inbred animal model to minimize immunologic variability. Complement was depleted by cobra venom factor, and activity in serum was monitored with a phagocytic assay. Bacterial specific antibodies were examined by an indirect radioimmunoassay, and animals with high levels of activity were excluded from anaysis. 4 h after aerosolization with Streptococcus pneumoniae, complement-depleted animals had cleared only 75% of the initial number of organisms, whereas saline-treated controls cleared 91% (P less than 0.01). Aerosolization with Pseudomonas aeruginosa was followed at 4 h by a twofold greater growth of organisms in the complement-depleted animals (446% of original deposition) as compared to the saline-treated controls (211% of original deposition) (P less than 0.02). Clearance of Klebsiella pneumoniae and Staphylococcus aureus were similar in complement-depleted animals and saline-treated controls. These experiments suggest that hypocomplementemia predisposes to bacterial pneumonia and may explain the high incidence of pulmonary infections in patients having impaired complement activity. Our results further indicate that varying defense mechanisms may be involved with clearing the lung of differing bacterial species.


Effects of alpha adrenergic blockade upon coronary hemodynamics. The effect of alpha adrenergic block-ade on coronary blood flow regulation at rest was studied in 11 normally innervated patients and 8 cardiac allograft recipients by measuring arterial pressure and coronary sinus blood flow by thermodilution before and after alpha adrenergic blockade with phentolamine. Coronary vascular resistance was calculated by using coronary sinus blood flow and mean arterial pressure, and metabolic demand was estimated by the product of systolic arterial pressure and heart rate. In addition, the coronary sinus blood flow response to tachycardia was examined in 9 innervated patients and 12 denervated patients, with measurements repeated after phentolamine in 8 of the 9 innvervated patients and 6 of the 12 denervated patients. There was a 7.3+/-4.4% increase in coronary sinus blood flow in the innervated patients in response to alpha blockade, whereas the transplanted patients had an 8.2+/-1.8% fall in coronary sinus blood flow, despite equivalent changes in rate pressure product. The innervated patients also demonstrated a significantly greater increase in coronary sinus blood flow than did the transplanted patients during the first 5 s of an abrupt increase in heart rate (26+/-4 vs. 8+/-2.5 ml/min, P <0.001). This early response was blunted after alpha adrenergic blockade. We conclude that there is basal alpha adrenergic tone present on the coronary vasculature in man that is withdrawn by a sudden increase in heart rate.


gamma-Glutamyl transferase isoenzymes in human bile. The gamma-glutamyl transferase isoenzymes of bile were studied using electrophoretic, gel filtration, and ultracentrifugation techniques. In view of the known association of other biliary enzymes with lipids the effects of butanol extraction were investigated. The results show the presence of four isoenzymes of gamma-glutamyl transferase in bile, differing in electrophoretic mobilities, molecular size, and density. The correlation between the properties of biliary gamma-glutamyl transferase and of alkaline phosphatase is discussed.


Plasma levels of clobazam after three oral dosage forms in health subjects. As can be seen from the tables, the terminal half-life of clobazam is about 50 hours, and from a solid dosage form the peak plasma level occurs approximately 1.5 hours after ingestion. Thus, there is a significant, yet relatively short, dosage form delay effect when the solid dosage forms are compared to the rapidly available solution of the drug. However, based on the areas under the curve, comparison of the solid dosage forms with the solution indicates that the fraction of clobazam absorbed is 1. Pupil diameter measurement at 2, 4, and 6 hours after ingestion of clobazam correlated well with the plasma levels at these times. Pupils were constricted to the highest degree at 2 hours and approached the initial pupillary diameter at the 6-hour measurement.


Clobazam versus diazepam--a double-blind study in anxiety neurosis. Clobazam, a new antianxiety compound, was compared in a double-blind study  with diazepam in 40 neurotic outpatients. Twenty-three patients completed the trial under clobazam conditions while 17 patients completed the trial under diazepam conditions. The trial was conducted for a period of four weeks of active drug administration followed by a one-week period of placebo administration. Clobazam was administered in three divided doses of 30 to 40 mg/day, while diazepam was administered in three divided doses of 15 to 20 mg/day, following a fixed dosage schedule. No significant differences were noted between the two treatment conditions during the drug trial period. The patients on clobazam maintained greater improvement during the placebo trial period for the variables "somatic anxiety" and "nights of sleep disturbance." Simultaneous motor coordination tests (hand steadiness test) showed greater improvement on clobazam throughout the trial period in patients with an initial error score greater than 50 points. This difference was significant during the second week of the trial.


Lactostrepcins--acid bacteriocins produced by lactic streptococci. All 47 non-nisin producing strains of Streptococcus lactis and 12/13 strains of Str. lactis subsp. diacetylactis examined produced bacteriocins, for which the term lactostrepcins is suggested. Seven strains of Str. cremoris examined produced no bacteriocins active against 3 lactic streptococci strains used as indicators. The strains examined were divided into 3 groups: I, those producing lactostrepcins active against only one streptomycin resistant mutant of Str. lactis 60 indicator strain; II, those producing lactostrepcins active against all 3 indicator strains; III, those not producing lactostrepcins active against the indicator strains employed. The lactostrepcins were sensitive to various proteolytic enzymes and to phospholipase D, but retained full or partial activity after dialysis. Most of the bacteriocins studied were fully active only within the pH range 4.2--5.0 and were reversibly inactivated at pH 7.0 or 8.0. Results suggested occurrence of 4 different lactostrepcins. The lactostrepcins produced by all group I strains were the same, but there were differences among the lactostrepcins produced by group II strains. Lactostrepcins killed some beta-haemolytic streptococci and some strains of Lactobacillus helveticus. One of the lactostrepcins was also active against certain Leuconostoc strains, but not against other Leuconostoc strains, nor against L. helveticus or other Gram-positive bacteria.


Blood and tissue distribution of gamma glutamyl transferase in calves. In five male and five female calves, we studied the tissue distribution of gamma-glutamyl transferase. The enzyme was mainly in kidney, pancreas, and liver; there was no sex-related difference. The relative hepatic and pancreatic specificity of the enzyme indicated that the measure of its activity in serum could be a test of hepatic or pancreatic damage in the calf. Serum activity measured within 159 samples of apparently healthy calves was 15.3 +/- 3.7 U/liter, not differing significantly from that of adult cow.


Nutrition policy--from neglect and uncertainty to debate and action. Until recent years, nutrition has received little attention in U.S. agriculture, food, and health policies. This circumstance is changing. In the late sixties and early seventies, reports of hunger and malnutrition sparked public reaction and a shift in policy. The White House Conference on Food, Nutrition and Health in 1971 prompted the Senate select Committee on Nutrition and Human Needs to address itself to this problem, with the result that the Food Stamp, child nutrition, WIC, and Nutrition for the Elderly programs were initiated or expanded. Then, in the mid-seventies, the Select Committee turned its attention to broader issues of nutrition and health and declared that the goal of any food system is the maintenance and improvement of nutritional health of the population. This objective emerged as public policy in the Food and Agriculture Act of 1977. As the Select Committee continued its work, problems of overnutrition became more apparent. The culmination of its studies was the issuance early in 1977 of the "Dietary Goals for the United States," designed to improve the nutrition and reduce health problems of the population. To that same end, the Select Committee has also made recommendations regarding food labeling and nutrition education.


Comparison of oxazepam, flurazepam and chloral hydrate as hypnotic sedatives in geriatric patients. In a four-week study, a comparison was made of oxazepam, flurazepam and chloral hydrate as hypnotic sedatives in 17 geriatric patients. Each drug was given alone for six nights, with a two-night placebo interval following each phase. Each patient completed an additional placebo phase (up to six nights) before each drug phase. The number of awakenings per night and the sleep latency (time required to fall asleep) were determined from the patients' reports and from the reports of a nurse-observer. Only for oxazepam was the number of patient-reported awakenings per night significantly less than for placebo, although with both oxazepam and flurazepam the awakenings were fewer than with chloral hydrate. According to the patient-reports, sleep latency was significantly lower with flurazepam than with placebo; for oxazepam and chloral hydrate the latencies were not significantly different from those for flurazepam or placebo. Only for oxazepam were the patients' ratings of sleep quality significantly greater than for placebo. The objective assessment of sleep by the nurse-observer usually confirmed the patients' assessments. Morning drowsiness was the most common side effect, reported equally for placebo and for the active drugs. Drowsiness during the day was reported less frequently for oxazepam than for flurazepam, chloral hydrate or placebo. It is concluded that oxazepam is safe and efficacious for the short-term management of insomnia in the elderly.


Antibodies of restricted heterogeneity directed against the cardiac glycoside digoxin. Intravenous injection of New Zealand White rabbits with type III pneumococcal polysaccharide vaccine conjugated with the cardiac glycoside digoxin resulted in the production of both antidigoxin and anti-type III pneumococcal polysacharide antibodies. Among antisera of 12 rabbits examined during their peak antibody production periods, 1 to 20 mg (mean, 5.4 mg) of antidigoxin antibody could be recovered from 1 ml of serum. Antisera from five of these 12 rabbits contained antidigoxin antibodies of restricted heterogeneity as demonstrated by urea-polyacrylamide disc gel electrophoresis of fully reduced and alkylated antibodies. From the antisera of four of these five rabbits, electrophoretically homogeneous antibodies (1 to 5 mg/ml antiserum) could be isolated by affinity chromatography on ouabain-amine-Sepharose columns. The structural homogeneity of two of these antidigoxin antibodies was confirmed by amino acid sequence analysis of purified light chains through the first hypervariable region. These data suggest that the conjugation of small molecules to bacterial polysaccharide vaccines may provide a general method for synthesis of immunogens that can regularly elicit antihapten antibodies of restricted heterogeneity.


Toxicity of Na251CrO4 when used to label rat lymphocytes. Lymphocytes preincubated with various activities of Na251CrO4 were tested for their ability to induce a local, popliteal lymph node graft-versus-host (GVH) reaction. Incubation with 100 micronCi of the isotope per ml of cell suspension impaired the GVH activity, 25 micronCi gave a reduced response in 1 out of 3 experiments, whereas 10 micronCi had no effect. The reduced GVH response was due to radiation rather than chemical damage of the cells. Preincubation of the cells with either cold Na2CrO4 or outdated Na251CrO4 with concentrations of Cr identical to the ones used to label cells with 100 micronCi/ml cells did not impair the GVH reactivity. The cellular uptake of Na251CrO4 showed first-order kinetics over wide concentration ranges. Mitomycin C-treated lymphocytes showed a more heavily depressed GVH reactivity than would be expected from their moderately reduced ability to accumulate in the lymph nodes. It is recommended that doses of Na251CrO4 in excess of 10 micronCi/ml cells should not be used for functional studies of lymphocytes.


Separation and characterization of anti-benzylpenicilloyl (BPO) antibodies. I. Biochemical and biophysical properties of anti-BPO-IgG obtained by affinity and subsequent ion-exchange chromatography. Anti-BPO antibodies were purified by means of affinity chromatography using AH-Sepharose 4B coated with covalently bound BPO groups. Specific elution was achieved by the hapten analogue BPO-epsilon-aminocaproic acid (BPO-EACA); desorption of the remaining antibody was performed thereafter by 0.1 M acetic acid. The resulting antibody fractions--hapten-eluted antibody (H-Ab) and acid eluted antibody (A-Ab), respectively--were further separated by ion-exchange chromatography which led to the appearance of 3 subfractions in the case of H-Ab (H1, H2, H3) and 2 subfractions in the case of A-Ab (A1 and A2). In liquid isoelectrofocusing an inhomogeneous pattern resulted. The bulk of antibodies focused between pH 6.5 and 7.0. The average avidity of H-Ab was found to be higher than that of A-Ab suggesting that avidity may influence the elution pattern in affinity chromatography. The hydrophobic influence of the "spacer" and/or interactions of antibodies directed against the hydrophobic regions of the BPO group may explain why a considerable part of the antibodies could be recovered from the immunosorbent only by acid elution.


The indirect assay for leukocyte migration inhibitory factor (LIF)--standardization and the effect of pH. In the indirect assay for leukocyte migration inhibitory factor (LIF), lymphokine-rich supernatants were obtained by culture of stimulated lymphocytes and then tested for LIF activity in agarose plates using purified granulocytes as target cells. Studies on the standardization of the conditions under which LIF acts on the target cells are described, with emphasis on the use of "standard" supernatants of known LIF activity and the influence of pH on the action of LIF and the sensitivity of the assay. The observation that LIF activity is reduced when the ambient pH falls below 7.2 is suggested as an explanation firstly for the "escape" phenomenon seen particularly in capillary tube assays for LIF, and secondly for the reduced sensitivity of the capillarly tube assay in comparison with the corresponding agarose plate assay.


Studies on the characterization of isolated renin-containing granules: the storage form of renin. Renin-containing granules isolated by isopycnic zonal centrifugation were partially characterized biochemically. Extracts of the granules were found to have a dual pH optimum at 5.5-6.0 and 7.0-7.5 and possess linear time-dependence of product formation when reacted with homologous renin substrate. Extracts also possessed a first order reaction constant of 0.713 X 10(-2). Treatment of the extracts with ammonium sulfate increased the velocity of the renin/renin substrate reaction though it continued to be linear with respect to time. This partially purified renin preparation was found to have a Km=3.9 X 10(3) ng/ml. Gel filtration of this preparation of renin demonstrated the presence of an active protein with renin activity and a molecular weight of 59,000 daltons in addition to an inactive protein of molecular weight 13, 750 daltons which may be a potential inhibitor of the renin/renin substrate reaction. The former protein or "big renin" may be the storage form or porhormone state of renin within the renin-containing granules of the juxtaglomerular cells.


Enrichment of in vitro and in vivo immunologic activity of purified fractions of calf thymic hormone. Thymus humoral factor (THF), a thymus hormone which participates in the processes leading to acquisition of immunocompetence of lymphoid cells has been isolated in our laboratory by a stepwise gel filtration through various Sephadex columns. THF so isolated appears to be a polypeptide of 3,000 mol wt which contains approximately 30 amino acid residues. Here we have tested the biological activity of THF fractions of successive degrees of purity upon lymphoid cells from both intact and neonatally thymectomized mice. The lymphoid cell populations were treated with the various THF fractions by in vitro incubation for a short time and by repeated injection in vivo. The treated cells evidenced increased ability to react in the graft-versus-host assay in vivo and in mixed lymphocyte cultures in vitro concomitantly with the rise of intracellular cAMP. On the other hand no activity whatsoever was shown by any of the control materials tested. These bioassays permitted isolation of fractions progressively more active than the original crude dialyzate of thymus extract tested. Thus the active peptide component of THF eluted from DEAE Sephadex A-25 column was estimated to be 2 X 10(4)-fold more active than the crude dialyzate of thymus extract which served as a starting material.


Determination of perazine serum levels by gas liquid chromatography under clinical routine conditions. A quantitative gas liquid chromatographic method for the determination of serum levels of perazine (10-[3'-(1''-methyl-4''-piperazinyl)-propyl]-phenothiazine) is described. Perazine is used as a neuroleptic drug. The main problem consists in optimizing the chromatographic system. A sensitivity of appr. 60--150 nmol/1 (20--50 microgram/1) serum is achieved. Examples of optimization, analyses with patient samples, and the reproducibility of the results are presented.


Increased urinary excretion of keratan sulfate in fucosidosis. In two children exhibiting the clinical symptoms of fucosidosis, the diagnosis was biochemically ascertained by the demonstration of a profound altpha-L-fucosidase deficiency in cultured skin fibroblasts. The non-dialysed urines of these fucosidosis patients were separated into two fractions by chromatography on Biogel P-2. The first fraction containing the glycosaminoglycans was further fractionated on Dowex 1 X 2 by stepwise elution with increasing NaCl concentrations. Keratan sulfate-chondroitin sulfates attached to the same peptide core were assayed and characterised mainly in the fractions eluted with 1.25, 1.5, 2.0 and 3.0 mol/1 NaCl. Whereas the excretion of normal children of the same age was found to be 0.77 mumol glucosamine equivalents per day in the 2 mol/1 and 3 mol/1 NaCl fraction, the two patients excreted 6.7 (M. C.) and 3.5 (M. S.) mumol glucosamine equivalents per day, respectively. Since keratan sulfate contains alpha-fucose at the non-reducing terminal, this increase in excretion of long chain keratan sulfate in fucosidosis could result from impaired degradation of keratan sulfate, due to the alpha-fucosidase deficiency.


Comments on "Studies of the interaction of psychological and pharmacological determinants of smoking" by Schachter et al. The work by Schachter and his colleagues on the psychological and pharmacological determinants of smoking is reviewed. The experiemental procedures are critically examined, and some are judged to be in violation of ethical standards for the conduct of research with human participants. Questions are raised concerning matters of informed consent, the description of debriefing procedures, and use of the college classroom as a psychological laboratory.


Purification and properties of glutamate dehydrogenase from the mantle muscle of the squid, Loligo pealeii. Role of the enzyme in energy production from amino acids. 1. The activity of glutamate dehydrogenase was measured in the tissues of the squid, Loligo pealeii. The enzyme occurs in high activity in digestive pouch, systemic heart, and all muscle tissues. 2. Glutamate dehydrogenase from mantle muscle is located intra-mitochondrially, has a molecular weight of 310,000, and is electrophoretically similar to the enzyme from all other squid tissues. 3. The enzyme from mantle muscle  was purified 40-fold by elution from DEAE-cellulose and used for kinetic studies. The enzyme is NAD+-specific, activated by ADP, AMP, and leucine, and inhibited by GTP, GDP, ATP, and reaction products (in particular NADH). 4. Squid glutamate dehydrogenase shows an almost absolute dependence on ADP. The purified enzyme is activated over 100-fold by saturating concentrations of ADP (Ka = 0,75 7M); The pH optima are also altered significantly by ADP. 5. The enzyme appears to be kinetically adapted to favour glutamate oxidation in comparison to glutamate dehydrogenase from other resources. The evidence indicates that the primary role of glutamate dehydrogenase in squid mantle muscle is in regulating the catabolism of amino acids for energy production.


Ionic blockage of the light-regulated sodium channels in isolated rod outer segments. We have investigated, with osmotic techniques, the light-regulated Na+ channels in rod outer segments (ROS) and ROS fragments freshly isolated from the frog retina. Values of Na+ permeability (PNa) similar to those observed electrophysiologically in the retina were observed using the osmotic technique (continuous flow) described by Korenbrot and Cone. In the other osmotic techniques that we explored, PNa was greatly diminished, if not completely suppressed; however, we found with these techniques that antioxidant conditions (N2 atmosphere or EDTA) significantly increased PNa, suggesting that the Na+ channels are highly sensitivive to membrane oxidation. Using the continuous flow technique, we investigated the H+ and Ca++ dependence of the Na+ channels and found that both of these ions, at micromolar activities, can block the channels. Raising the external H+ activity decreases PNa (reversibly) in a single "sigmoidal" response with an apparent pKa of 5.8. Similarly, in the presence of the ionophores X537A or A23187 which allow equilibration of Ca++ across membranes, the Na+ channels are blocked when the external Ca++ activity is increased from 10(-7) to 10(-5) M. This high sensitivity to both H+ and Ca++ ions suggests that high field strength anionic sites may exist in or near the Na+ channels and that the channels are blocked when these sites bind H+ or Ca++ ions.


Nitrogen regulation of glutamine synthetase in Neurospora crassa. A higher activity of glutamine synthetase (EC 6.3.1.2) was found in Neurospora crassa when NH4+ was limiting as nitrogen source than when glutamate was limiting. When glutamate, glutamine or NH4+ were in excess, a lower activity was found. Immunological titration and sucrose gradient sedimentation of the enzyme established that under all these conditions enzyme activity corresponded to enzyme concentration and that the octamer was the predominant oligomeric form. When N. crassa was shifted from nitrogen-limiting substrates to excess product as nitrogen source, the concentration of glutamine synthetase was adjusted with kinetics that closely followed dilution by growth. When grown on limiting amounts of glutamate, a lower oligomer was present in addition to the octameric form of the enzyme. When the culture was shifted to excess NH4+, glutamine accululated at a high rate; nevertheless, there was only a slow decrease in enzyme activity and no modification of the oligomeric pattern.


Biochemical and biophysical characterization of light and heavy density hepatitis A virus particles: evidence HAV is an RNA virus. Heavy density HAV was also shown to be sensitive to low concentrations of RNase. The results of these biophysical and biochemical studies strongly support the notion HAV is an enterovirus.


Behavioral rhythms in schizophrenia.

Behavioral rhythms in schizophrenia. Daily behavioral observations were made for several years on 10 male schizophrenic patients and on three male patients with organic brain disorders. Analysis of these data showed strong cyclic components in the five schizophrenic patients with predominantly hebephrenic symptomatology. Period lengths noted were about 2 days, 5 to 6 days, 30 days, and a longer cycle of 40 to 100 days duration. Antipsychotic medications appear to have a suppressant effect, but tricyclic antidepressants may enhance pre-existing rhythms.


Clinical course of adult metachromatic leukodystrophy presenting as schizophrenia. A report of two living cases in siblings. Adult metachromatic leukodystrophy is a demyelinating disease due to an inherited lack of arylsulfatase A activity. The purpose of this paper is to present the characteristics of this disorder as they occurred chronologically in two siblings, prior to and subsequent to the appearance of gross neurological deficits. A deficit in spatial relationships, as contrasted with verbal abilities, was observed initially in both cases at age 13. Initial psychiatric symptoms were noted at age 16 and 18, with both patients being diagnosed subsequently as schizophrenic. Gross neurological deficits were observed 2 and 13 years, respectively, after the appearance of psychiatric symptoms. A deficit in spatial relationships may be a very sensitive early indicator of adult metachromatic leukodystrophy.


Neurobiology of Polyorchis. I. Function of effector systems. The electrical correlates of activity in the effector systems responsible for swimming, crumpling and postural changes have been recorded in the anthomedusan Polyorchis penicillatus. Motor spikes (pre-swim pulses), that initiate swimming contractions, appear without delay at distant sites on the inner nerve-ring in unstimulated preparations. Levels of Mg++ anaesthesia which block the neuromuscular junctions between PSP giant neurons and swimming muscle do not affect PSP activity. Swimming muscle potentials can be recorded from subumbrella and velar muscle sheets using extra- and intracellular electrodes. These action potentials have a distinct plateau and are propagated in a myoid fashion. Resting potentials average -70 mV with spikes overshooting zero by some 62 mV. The effects of repetitive stimulation are described. Extracellular recordings indicate that neuronal pathways may play a major role in mediating crumpling, unlike many other species where epithelial pathways are more important. Endodermal spikes recorded intracellularly from the radial and ring canals have amplitudes of some 92 mV arising from resting potentials that average -55 mV. Repetitive stimulation causes a decrease in amplitude and increase in duration of epithelial action potentials. Tentacle length is controlled by a pacemaker system located in both nerve rings. The frequency of spikes (PTPs) generated by this system determines the length and tonus of tentacles. The neuromuscular junctions between the motor neurons and tentacle muscle are Mg++ sensitive and show facilitating properties.


Cytochemical demonstration of adenylate and guanylate cyclases in vascular smooth muscle of circle of Willis. The cytochemical localization of adenylate cyclase and guanylate cyclase was studied in the arteries of the circle of Willis in dogs. The reaction products of both adenylate and guanylate cyclases were similarly distributed and selectively localized predominantly adjacent to sarcoplasmic reticulum and sparsely to mitochondria and outer nuclear membranes of vascular smooth muscles. The observations could suggest a close association of the intracellular localizations of both cyclases and the intracellular calcium storage sites, and ultimately contribute to our complete understanding of regulation of cerebral blood flow and vasospasm.


Abnormal cellular copper metabolism in the blotchy mouse. Defective copper metabolism was demonstrated in male mice bearing the blotchy (Moblo/y) allele at the mottled locus on the X-chromosome. Copper absorption from the gut was only 64% of that found in normal mice and hepatic copper levels were only 56% of the controls. Ceruloplasmin and heart cytochrome c oxidase activities were normal, yet lysyl oxidase activity from cultured fibroblasts was only 45% of control levels. Copper accumulated in fibroblasts cultured from these mutants to values that were five times normal. The accumulation of copper in the fibroblasts was associated with a protein of approximately 12,000 molecular weight.


Loss of in vitro inactivation of rat liver tyrosine aminotransferase with dietary vitamin B6 restriction. Tyrosine aminotransferase, in the presence of 8 mM L-cysteine, is inactivated in incubated liver homogenates prepared from normal rats, but not in those from rats deprived of vitamin B6. In this study we fed rats a diet deficient in vitamin B6 to determine the length of time required for in vitro inactivating activity to be lost from liver homogenates. After 2 weeks, the half-life of tyrosine aminotransferase in liver homogenates from vitamin B6-deficient rats was 5.9 hours, and from control rats, 1.8 hours. After 3 weeks, tyrosine aminotransferase was no longer inactivated in homogenates prepared from livers of deficient rats. The pyridoxal phosphate (PLP) concentration of plasma from rats fed the vitamin B6-deficient diet dropped from 89 ng/ml to 14 ng/ml after 1 week and to 7 ng/ml after 2 weeks. In 5 weeks the PLP concentration of liver from vitamin B6-adequate rats increased from 2.9 microgram/g to 6.6 microgram/g while in deficient rats it dropped to 2 microgram/g. The loss of tyrosine aminotransferase inactivating activity in the livers of vitamin B6-deficient rats occurred at approximately the same time that the concentration of PLP in the livers of rats fed the two diets began to show marked differences.


Characterization of thiamine diphosphatase in rat small intestine. The properties of thiamine diphosphatase (TDPase) and p-nitrophenylphosphatase (p-NPPase) in rat small intestine were investigated. TDPase activity, like p-NPPase activity, was high in the mucosa and in the proximal region. Both activities were high in the membrane-associated fractions of the duodenal mucosa. Furthermore, TDPase had the same properties as intestinal alkaline phosphatase (al-Pase). These results suggest that thiamine diphosphate (TDP) and p-nitrophenylphosphate (p-NPP) are hydrolyzed by a single enzyme, al-Pase, in the intestine.


The isolation and characterization of a trypsin inhibitor from Kintoki bean (Phaseolus vulgaris). A trypsin inhibitor was isolated from beans of Phaseolus vulgaris, cultivar. Kintoki, and the specific activity increased 200 times as high as that of the crude extract. It was homogeneous on several electrophoreses and the molecular weight was about 13,000. The amino acid composition was characterized by high ratios of cystine, aspartic acid, and serine. It inhibited trypsin in a molar ratio of 1 : 1 and alpha-chymotrypsin in a molar ratio of 2 : 1. It, however, inhibited neither pepsin nor pronase. It was relatively stable to heat treatment in the acidic medium, but not in the alkaline medium. Neither pepsin nor pronase destroyed the inhibitory function.


Effects of various metabolites (sugars, carboxylic acids and alcohols) on riboflavin formation in non-growing cells of Ashbya gossypii. The effects of various sugars and sugar derivatives on flavinogenesis were examined using non-growing cells of a high flavinogenic mold, Ashbya gossypii. Glucose, fructose and galactose were found to be the most stimulative. Glycerol and glucono-delta-lactone were less stimulative; next in order were n-propanol, n-butanol, glycols and butanediols, which were likewise effective; acetate, lactate and pyruvate were slightly stimulative. In contrast, ribose, xylose, arabinose, ribitol, citrate, succinate, oxaloacetate, glyoxylate and malate were rather inhibitory, in additions at 1.0%. Among these compounds, ethanol (1%) greatly stimulated  riboflavin formation. Maximum flavinogenesis with the above stimulants was attained by the additions of 1% ethanol, 1.25--3.0% glucose, 1.25% glycerol, 4.0--6.0% propane and butanediols, 1.0% pyruvate and 0.9% acetate after 37 hr incubation, respectively. These compounds inhibited flavinogenesis with increasing concentrations above their optimum concentrations. The stimulation effect of ethanol far exceeded those of other stimulants but ethanol had almost no effect on growth and pH values during incubation. With the addition of ethanol (1%) during incubation, maximum formation (1,776 microgram/g wet mycelia) of riboflavin was achieved when added at the start of incubation and the most effective utilization was observed when added at the logarithmic phase of flavinogenesis, although the maximum formation of riboflavin in the latter case was much lower than in the former case. The relation of sugar metabolism, especially ethanol metabolism, to flavinogenesis was discussed with the flavinogenic activities of these additives.


Multiple-scored tablets. Weight and content uniformity of subdivisions and distribution of active constituent within and between tablets. Using three brands of multiple-scored levodopa tablets, B.P. (500 mg) and one brand of sulphamethoxypyridazine tablets B.P. (500 mg) the weight and content uniformity of the subdivisions has been examined. It is shown that quartering of such tablets can result in subunits which do not conform to recognized standards of weight uniformity, and in some instances content uniformity may be questionable. The homogeneity of distribution of active constituent between tablets has been determined and compared with that within tablets (between quarters of individual tablets). Statistical evaluation of the results is presented.


The effect of repeated administration of diftalone on enzyme induction. Plasma concentrations of diftalone have been examined in normal volunteers after a single dose (500 mg) and after 500 mg doses given twice daily for one week. An increase in post dosing urinary excretion of D-glucaric acid showed a correlation with the ratio of calculated to observed areas under the plasma concentration, time curve following the final dose in the multiple dosing studies, indicating that hepatic microsomal enzymes are induced after repeated administration of the drug. Single dose studies in the presence of aluminium hydroxide and sodium bicarbonate showed that the antacids had no significant effect on the absorption of diftalone.


The determination of sulphoxide in degraded phenothiazine formulations by difference spectrophotometry. A method is described for the assay of phenothiazine sulphoxides which may be formed in phenothiazine formulations during storage under unfavourable conditions. The assay is based upon the measurement of difference absorbance of the sulphoxide solution in 0.2 M hydrochloric acid relative to an equimolar solution reduced with zinc dust. A solvent extraction procedure avoids interference from colouring agents and coloured photolytic breakdown products of the phenothiazines. The assay is specific in the presence of intact drug, sulphone and co-formulated drugs for the formulations examined and is sensitive to 0.5% of the total phenothiazine present as sulphoxide. Many aqueous formulations stored in partially filled containers have been shown to contain up to 31.5% of the total phenothiazine as sulphoxide.


Species differences in the in vitro metabolic reduction of the amphetamine metabolite, 1-phenyl-2-propanone. Marked differences were observed, in the ability of fortified 9000 g liver homogenate supernatants from three species to reduce 1-phenyl-2-propanone to the corresponding alcohol. This metabolic keto-reduction was negligible in homogenates from the rat and extensive in the rabbit; guinea-pig liver homogenates had intermediate ability. Metabolic oxidation of 1-phenyl-2-propanol was negligible in all three species. The amount of deamination of amphetamine and of N-n-propylamphetamine was approximately equal, in vitro, in rats and guinea-pigs but two to three times greater in liver homogenates from rabbits. Approximately three times more deaminated products were formed from the in vitro metabolism of N-n-propylamphetamine than from amphetamine metabolism by all three species.


Effectiveness of pyridostigmine in reversing neuromuscular blockade produced by soman. The effects of pyridostigmine pretreatment on the neuromuscular blockade produced by soman in anaesthetized, atropinized animals have been studied on the soleus and anterior tibialis muscle (rhesus monkeys, cats and rabbits) and the gastrocnemius muscle (guinea-pigs and rats). Pyridostigmine pretreatment produced a complete recovery of neuromuscular function following blockade by soman; the rate of recovery was similar in all the species, suggesting a common mechanism of action. In the absence of pyridostigmine or if pyridostigmine was delayed until after blockade by soman, there was no recovery of neuromuscular function. Detailed studies in the guinea-pig showed that the recovery of neuromuscular function was related to the dose of soman and to the degree of carbamoylation of blood cholinesterase at the time of nerve agent challenge, i.e. to the dose of pyridostigmine and the time interval between the administration of pyridostigmine and soman. It is suggested that the effectiveness of pyridostigmine pretreatment is due to the carbamoylation of a portion of the tissue acetylcholinesterase, which protects it against irreversible inhibition by soman: after poisoning spontaneous decarbamoylation produces sufficient free acetylcholinesterase to restore normal function.


The relationship between cholinesterase inhibition in the chick biventer cervicis muscle and its sensitivity to exogenous acetylcholine. The effect of physostigmine has been studied on cholinesterase in homogenates of chick biventer cervicis muscles and on the contractile responses of the intact muscles to acetylcholine and carbachol. The concentration of physostigmine required to produce the maximum increase in sensitivity to acetylcholine almost completely inhibited the cholinesterase in muscle homogenates. This concentration of physostigmine had no effect on muscle contractures elicited by carbachol. By taking account of the combined effects of acetylcholine diffusion and enzymic hydrolysis, a quantitative theoretical relationship has been derived between the level of cholinesterase activity in cylindrical muscles and the fractional occupancy of the acetylcholine receptors in these muscles in the presence of different concentrations of exogenous acetylcholine. This theory attributes the thousand-fold increase in sensitivity to exogenous acetylcholine produced by anticholinesterases in chick biventer cervicis muscles largely to an alteration in acetylcholine concentration gradient within the muscle and accounts satisfactorily for the shift in the dose-response curve for acetylcholine which occurs after treatment of the muscles with various concentrations of physostigmine.


Pharmacological study of chicken airway smooth muscle. Isolated chicken bronchus contracts to carbachol, PGF2alpha, histamine, 5-HT, bradykinin and phenylephrine. The bronchial strips from the horse plasma sensitized domestic fowl contracted to specific sensitizing antigen in vitro (Schultz-Dale anaphylactic reaction) and were 5 to 10 fold more reactive to PGF2alpha, histamine and 5-HT compared with the bronchi from normal chickens. Second antigen challenge was inactive or produced a weak response (desensitization). Allowing the bronchi to rest for 1 h resulted in partial recovery of anaphylactic response. Bronchi which had partially contracted submaximally to carbachol, antigen or PGF2alpha, relaxed to isoprenaline, adrenaline and PGE1 and E2.


The evaluation of cardiovascular drugs in the anaesthetized, unrestrained rat. A method is reported which allows continuous long-term drug administration and simultaneous blood pressure measurement in the unanaesthetized unrestrained rat. The external jugular vein and abdominal aorta were cannulated and the opposite ends of the cannulae were passed subcutaneously and exteriorized at the back of the head. They were then passed through a spring attached at the lower end to the skull and, at the upper end, to a counter-weighted cantilever. In rats so prepared, infusion of angiotensin amide 200 ng kg-1 min-1 caused a rise of blood pressure which lasted the 48 h infusion period. Heart rate decreased initially but recovered within 6 h. Angiotensin amde 30 ng kg-1 min-1, infused up to seven days, was without effect on blood pressure or heart rate, and both doses of angiotensin amide failed to alter cardiac catecholamine turnover. Hydralazine, mecamylamine and clonidine reduced blood pressure to 63, 62 and 84% of control respectively while clonidine induced a transient increase before its depressor effect. Heart rate was increased by hydralazine to 138%, and decrease by clonidine to 74% of control, and was unaffected by mecamylamine. The magnitude of pressor response to noradrenaline, tyramine and angiotensin was reduced by hydralazine and increased by mecamylamine. Clonidine increased the pressor response to angiotensin but had no effect on that to noradrenaline or tyramine.


Physicochemical properties of amphoteric beta-lactam antibiotics I: Stability, solubility, and dissolution behavior of amino penicillins as a function of pH. The degradation rate, solubility, and dissolution rate of amino penicillins, amoxicillin, ampicillin, epicillin, and cyclacillin, were determined quantitatively as a function of pH. In the pH range studied, 0.30-10.50, the degradation of amoxicillin and epicillin followed pseudo-first-order kinetics to give the same type of pH-rate profiles as those of ampicillin and cyclacillin. Cyclacillin anhydrate was the most soluble, followed in order by ampicillin anhydrate, ampicillin trihydrate, amoxicillin trihydrate, and epicillin anhydrate. These pH-solubility profiles showed showed U-shaped curves. The dissolution rate constants from the rotating disk were analyzed by the simultaneous chemical reaction and diffusion models. Their relative bioavailability after a single oral administration was assessed from their physicochemical properties determined in vitro.


Mechanism of adsorption of clindamycin and tetracycline by montmorillonite. IR and X-ray analyses of the interaction of clindamycin with montmorillonite indicate that clindamycin is adsorbed by a cation-exchange mechanism under pH conditions favoring the cationic form of the drug and by physical adsorption when the unionized drug is present. This physical adsorption is relatively weak since the drug is readily desorbed by alkaline washing. Tetracycline is adsorbed by cation exchange at low pH values where the +00 species predominates. Complexation with divalent interlayer cations contributes significantly to adsorption at higher pH values where the +-0 and +-- species exist. In a strongly alkaline solution, the 0-- species was not adsorbed in the interlayer space of montmorillonite but rather produced an external calcium-tetracycline complex. This study illustrates the utility of X-ray and IR analyses in elucidating the mechanisms responsible for clay-drug interactions.


Oxidative degradation of pharmaceutically important phenothiazines III: Kinetics and mechanism of promethazine oxidation. The kinetics of the thermal degradation of promethazine in an acidic medium under various conditions were investigated. The degradation of promethazine and the formation of some degradation products were studied under aerobic and anaerobic conditions. The influence of pH, metal ions such as copper(II) and iron (III), and antioxidants was investigated. In an oxygen-saturated medium, promethazine generally followed first-order kinetics. Increasing the pH increased the degradation rate to a limiting value at pH 5. Addition copper (II) increased the degradation rate over the whole process, while iron (III) caused an increase for only a short time. Ascorbic acid sometimes increased the degradation rate, while higher concentrations of hydroquinone also accelerated the degradation. Pyrosulfite did not have any influence. Under anaerobic conditions, promethazine degraded only in the presence of copper (II) and iorn (III) ions. As a result of the studies on the qualitative and quantitative aspects of the oxidation process, a mechanism for the oxidative degradation of promethazine is suggested. Promethazine 5-oxide and a number of degradation products without intact side chains are formed via a semiquinone free radical. The influence of several factors on the degradation process is discussed.


Nature of amorphous aluminum hydroxycarbonate. The titration of sodium carbonate with aluminum nitrate is shown to produce amorphous aluminum hydroxycarbonate. This compound is not stoichiometric, although the maximum carbonate to aluminum ratio appears to be 0.5. The pH conditions for achieving the maximum carbonate content are concentration dependent. A model for the particle surface at the solution interface is proposed. This model accounts for the presence of carbonate directly coordinated to the aluminum and carbonate adsorbed by electrostatic forces. Sodium is present in the diffuse layer and is, therefore, not an integral part of the structure.


Interference in assays for hydralazine in humans by a major plasma metabolite, hydralazine pyruvic acid hydrazone. The present study showed that published spectrophotometric and GLC methods for hydralazine in plasma do not distinguish between the drug and a major plasma metabolite, hydralazine pyruvic acid hydrazone. These methods involve the acid treatment of the sample, which hydrolyzes that hydrazone back to hydralazine. A specific GLC assay for the hydrazone was developed and involves its selective extraction from plasma and transformation to 3-trifluoromethyl-s-triazolo[3,4-a]phthalazine. This derivative could be sensitively measured by GLC using an electron-capture detector. With this procedure, it was shown that most "apparent hydralazine" in plasma is the hydrazone, which forms rapidly from hydralazine and endogenous pyruvic acid. Previous work indicated that the hydrazone was inactive when administered intravenously to rabbits.


Automated potentiometric procedure for studying dissolution kinetics acidic drugs under sink conditions. An automated potentiometric procedure was used for studying in vitro dissolution kinetics of acidic drugs. Theoretical considerations indicated that the pH-stat method could be used to establish approximate sink conditions or, possibly, a perfect sink. Data obtained from dissolution studies using the pH-stat method were compared with data obtained from known sink and nonsink conditions. These comparisons indicated that the pH-stat method can be used to establish a sink condition for dissolution studies. The effective diffusion layer thicknesses for benzoic and salicylic acids dissolving in water were determined, and a theoretical dissolution rate was calculated utilizing these values. The close agreement between the experimental dissolution rates obtained under pH-stat conditions and theoretical dissolution rates indicated that perfect sink conditions were established under the experimental conditions used.


A controlled trial of antiemetics in abortion by PGF2alpha and laminaria. Women undergoing abortion by intraamniotic prostaglandin F2alpha were randomized to receive either prochlorperazine edisylate 10mg, hydroxyzine hydrochloride 100mg, or a placebo every four hours by intramuscular injection in a double-blind fashion. Vomiting was significantly more frequent in the placebo-treated group [0.2 +/- 1.5 SD episodes per patient, n=21]  than in the groups treated with prochlorperazine [1.2 +/- 0.5 episodes per patient, n=21] or hydroxyzine [0.3 +/- 0.8 episodes per patient, n=19]. The mean number of merperidine injections in the antiemetic-treated groups was lower than in the control group, but this effect was not statistically significant. There was no significant difference between the treated and the control groups in the interval from prostaglandin treatment to abortion.


Novel anxiolytic agents derived from alpha-amino-alpha-phenyl-o-tolyl-4H-triazoles and -imidazoles. Several new alpha-amino-alpha-phenyl-o-tolytriazoles and -imidazoles have been prepared in one step by means of a novel reductive rearrangement of the corresponding benzodiazepines with hydrazine hydrate. These new triazoles were found to have moderate sedative and muscle relaxing activity in mice (i.e., these compounds depressed the traction and dish reflexes at higher doses than did diazepam) but were very potent antagonists of the clonic convulsions induced in mice by the administration of pentylenetetrazole. Furthermore, they antagonized the lethality induced by thiosemicarbazide. While these new compounds were very active in mice, most were inactive in rats. These results are discussed with reference to the metabolism of compound 13.


Synthesis of N-hydroxyacetaminophen, a postulated toxic metabolite of acetaminophen, and its phenolic sulfate conjugate. The synthesis of N-hydroxyacetaminophen (N-acetyl-N-hydroxy-p-aminophenol, 4), a postulated toxic metabolite of acetaminophen (N-acetyl-p-aminophenol, 3), and its phenolic sulfate conjugate (potassium N-acetyl-N-hydroxy-p-aminophenyl sulfate) (13) is described. Potassium p-nitrophenyl sulfate was reduced to the hydroxylamine, acetylated, and treated with sulfatase to yield N-hydroxyacetaminophen. The structures assigned are supported by the spectral data (IR, UV, MS, 1H NMR, and 13C NMR). N-Hydroxyacetaminophen was found to be moderately unstable at physiological pH and temperature, whereas it phenolic sulfate conjugate was stable.


Neuroleptics related to butaclamol. Synthesis and some psychopharmacological effects of a series of 3-aryl analogues. The synthesis and some pharmacological effects of 16 3-aryl analogues of butaclamol, a new antipsychotic drug, are described. The animal models were predictive of neuroleptic activity as well as side effects commonly associated with neuroleptic therapy. The results indicate that the 3-substituent plays a critical role with regard to the potency of the compounds as well as to their tendencies to induce extrapyramidal side effects and/or hypotension.


5-Carboxamido-4-amino-3-isoxazolidone, and asparagine analogue. trans-Aziridine-2,3-dicarboxylic ester was used to prepare the required beta-chlorohydroxamic acid used in the synthesis of the title compound. The trans configuration of the asparagine analogue was established by hydrogenolysis to erythro-beta-hydroxyasparagine amide. Neither the title compound nor the intermediate aziridinehydroxamic acid (8) showed significant activity against the L1210 and P-388 tumors. The title compound was inactive as an inhibitor of asparagine synthetase from Novikoff hepatoma and did not inhibit the growth of some 25 bacteria and fungi.


Autosomal recessive postaxial polydactyly type A in a Sicilian family. Postaxial polydactyly type A was present in several members of a Sicilian family. The anomaly was probably transmitted as an autosomal recessive character. Two polydactylous subjects were also beta-thalassaemia carriers, but a linkage between the two mutant genes could be excluded. Two patients with hexadactyly had a fifth digital triradius.


The influence of various cations on the catalytic properties of clays. Polymerization of alanine adenylate in the presence of various clays in their Na form gave increasing degrees of polymerization in the following order: montmorillonite less than nontronite less than hectorite. With montmorillonite, presaturated with different cations the order was: Mg less Ca less than Fe less than Al less than Na. From all these clays, hectorite was the only one to enable also some polymerization of lysine.


Biotin. Its place in evolution.

Biotin. Its place in evolution. In the previous article we try to give an explanation for the success of models of enzymatic reactions in which coenzymes play a role. However, one vital cofactor that has thus far not yielded appreciably to this approach is biotin. We attempt to show that biotin presents a fundamentally new sort of problem to bioorganic chemistry; this problem requires consideration both of the origin of life and subsequent evolution in any attempt to understand the anomalies offered by this cofactor.


Covalent coupling of calf brain prolidase. Calf brain prolidase covalently bound to CNBr-Sepharose 4B, retained about 32% of the activity of the uncoupled enzyme. The free enzyme showed slightly greater stability than the bound preparation when stored at 20 degrees C or at 0 degrees C. However, in either case the free and bound enzymes were more stable at the lower temperature. Greater thermal stability was shown by the free enzyme than by the bound preparation over a temperature range of 25 degrees C-60 degrees C. The free and bound prolidase, with and without Mn+2, had maximal activity at pH 4.0. Although the bound enzyme showed a single maximum, the free preparation exhibited three pH maxima of 4.0, 9.0, and 6.5, in decreasing order of activity. The ions Ag+, Cu+, Hg+2, and Zn+2 were strongly inhibitory on the free enzyme, whereas inhibition of the bound enzyme, with the exception of Zn+2 , was less. Unlike the coupled enzyme, a stimulatory effect was obtained on the free preparation with Co+3, Mg+2, and Mn+2. Various other compounds were studied and their effects were noted.


Characterization of an alkaline protease associated with a granulosis virus of Plodia interpunctella. An alkaline protease was found to be associated with the granulosis virus of the Indian meal moth. Plodia interpunctella. The protease was located within the protein matrix of the occluded virus and hydrolyzed the major constituent of this matrix, a 28,000-dalton protein (granulin), to a mixture of polypeptides ranging in molecular weight from 10,000 to 27,000. A rapid, sensitive assay for the protease was developed using radioactively labeled granulosis virus as substrate. With this assay, the proteolytic activity could be detected by measuring the release of acid-soluble peptides from the labeled virus. The protease had a pH optimum of 10.5 and a temperature optimum of 40 degrees C and was inhibited by diisopropyl phosphorofluoridate, phenylmethylsulfonyl fluoride, and L-(1-tosylamido-2-phenyl) ethyl chloromethyl ketone. Purification of the protease from matrix protein was achieved by anion-exchange and gel permeation chromatography. The molecular weight of the isolated protease, determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and gel filtration, was approximately 14,000.


The urologic manifestations of XO/XY mosaicism. The mosaic karyotype XO/XY is expressed by a spectrum of genital phenotypes, ranging from normal male through ambiguous genitalia to normal female. The urologist may see some of these patients because of the following clinical presentations: 1) ambiguous genitalia with a chromatin-negative buccal smear, 2) hypospadias with cryptorchidism or 3) cryptorchidism with müllerian remnants, discovered unexpectedly during inguinal operations. Reliable karyotyping is mandatory for diagnosis of XO/XY patients. Management should include laparotomy with excision of any intra-abdominal gonads (testis and/or streak gonad) because these are prone to develop malignancies that may occur before puberty. Female sex assignment and a reconstructive operation are advised in cases with severely deficient virilization of the genitalia.


Bilateral trifid ureters associated with fused kidney, ureterovesical stenosis, left cryptorchidism and angioma of the bladder. The first case of bilateral trifid ureters associated with fused kidney, ureterovesical stenosis, a left undescended testicle and angioma of the bladder is reported. Bilateral ureteroneocystostomy and left orchiopexy were done. The patient is well 3 months postoperatively with no urinary tract infection and stable renal function.


[The determination of a small amount of chlorine in the atmosphere by using 4-nitroaniline (author's transl)]. The determination of small amount of chlorine in the atmosphere is carried on ordinarily by omicron-toluidine method at present. However, this method is not recommended because the issued colour is unstable and omicron-toluidine itself belongs in the first group member of the Ordinance on Prevention of Particular Chemical Hazards Substances. Authors recommend 4-nitroaniline method, reported by J. Gabbay etc., because this method is stable and more practical. Although the colour that issued from the reaction of chlorine with 4-nitroaniline is affected by the co-existence of reducing gas, e.g. SO2, H2S or organic gas, but this effect may be removed the co-existence of reducing gas, e.g. SO2, H2S or organic gas, but this effect may be removed by introducing the sample gas into the layer of H2SO4-52Cr2O7 mixture immersed in glassfiber filter for micro-analysis. In this paper, influence of pH of the issued colour, stability, stream velocity of samplegas and effect of reducing substance are discussed.


[Effect of adrenergic beta receptor blockaders on bronchial patency in ischemic heart disease]. The effect of beta-adrenergic blocking agents inderal, trazicor, visken, benzoral and aptin on the bronchial patency was studied by means of four ventilation indices in 85 patients, 62 of whome had various diseases of the lungs; 44 of these patients had bronchial obstruction. It was established that all beta-blocking agents are detrimental to bronchial patency in patients with concomitant bronchial obstruction. In exacerbation of bronchial asthma, beta-blocking agents are contraindictaed. When it is necessary to prescribe beta-blocking agents for other patients with concomitant bronchial obstruction visken is the drug of choice. It should be given per os, with preliminary control of the function of external respiration.


[Transient myopia with angle closure glaucoma. Case report and biometric findings (author's transl)]. A 36-year-old man suffered from diarrhoea for some weeks and was treated with Salazosulphapyridie and Fluocortulon when he developed a transient myopia combined with an angle closure glaucoma. The refraction of his mild hyperopic eyes (+0,5 dpt) changed to -5,5 dpt and the intraocular pressure increased to 40 mm Hg in the right and 42 mm Hg in the left eye. Accommodation and pupillary reflexes were normal. Ultrasonic examination showed an increased axial diameter and a forward shifting of the lens. The resulting flattening of the anterior chamber induced the angle closure glaucoma. All symptoms disappeared within a week after the administration of Fluocortulon had been discontinued.


Semisynthetic derivatives of pyrrolizidine alkaloids of pharmacodynamic importance: a review. Pyrrolizidine alkaloids occur in more than 40 genera. Among these are two important genera, Senecio and Crotalaria, which have been responsible for heavy losses of livestock and poisoning in man due to their hepatotoxicity. Pyrrolizidine alkaloids are also reported to possess a number of other biological activities. Attempts have been made to convert the pyrrolizidine alkaloids to new structures by synthetic modifications. One hundred and twenty semisynthetic compounds were developed and investigated pharmacologically. The range of activity exhibited was hypotensive, local anesthetic, ganglion blocking, neuromuscular blocking and antispasmodic. The five most promising compounds of these series were subjected to detailed pharmacological investigations.


An adverse effect of insulin on the oxygen-release capacity of red blood cells in nonacidotic diabetics. Oxyhemoglobin dissociation curves (ODC) were performed on blood from newly diagnosed, nonketotic diabetics prior to and following initial insulin treatment and from ambulatory juvenile diabetics before and after their usual morning insulin. In 10 newly discovered diabetics the average P50 at in vivo pH was normal prior to insulin (26.2 mm Hg), decreased to 24.5 mm Hg (p less than 0.005) on the day following the initial insulin administration, and was within normal limits (26.9 mm Hg) when the diabetes was finally well controlled and red cell 2,3-diphosphoglycerate (2,3-DPG) had risen to elevated levels. Oxygen affinity of hemoglobin was closely correlated with the content of red cell 2,3-DPG (r = 0.61, p less than 0.001) but was unrelated to the level of hemoglobin Alc. In 40 juvenile patients the average P50 was also normal prior to insulin administration but was significantly lower 3-4 hr after they had received their usual insulin dose (p less than 0.001). The study indicates that insulin administration to diabetics with high blood glucose levels may lead to transient decreases in red cell 2,3-DPG and in oxygen-releasing capacity of the red blood cells.


Kinetic properties of pyruvate kinase of Neurospora crassa at physiological pH. The kinetic properties of pyruvate kinase (EC 2.7.1.40) extracted from the mycelia of Neurospora crassa were examined at physiological pH to determine the role of the enzyme in the regulation of glycolysis. The velocity curve with the substrates, phosphoenolpyruvate and adenosine diphosphate, are hyperbolic. The effect of magnesium, potassium, or calcium on the enzyme is influenced by the pH but not to the extent that would change their role as cofactor or inhibitor. Adenosine triphosphate and citrate remain strong inhibitors even with changes in pH. Fructose-1,6-diphosphate and glucose-6-phosphate are the dual positive effectors at physiological pH. Valine is the only amino acid that inhibits the enzyme at a concentration range of valine found in the mycelial juice. Thus, the properties of the enzyme at physiological pH are significantly different from those observed at neutral pH of the usual assay conditions, but its role as a key regulator of glycolysis is unchanged.


The influence of pH on the growth rate and viability of neutrophilic and acidophilic streptomycetes. The influence of pH on the specific growth rates of two acidophilic and two neutrophilic soil streptomycetes was studied. The acidophiles had maximum growth rates over a broad range from pH 4.5 to 5.5, while the neutrophiles had clearly defined optima at pH 7.0. Mycelium of neutrophiles was less tolerant of acidity than that of acidophiles; both showed decreased viability at pH levels below those which allowed growth. Spores of neutrophiles and acidophiles were equally tolerant of acidity and this may allow the former to survive in acid soils. Both spores and mycelium of acidophiles remained viable at pH levels above those allowing growth.


Cell wall autolysis in log phase cells of Micrococcus lysodeikticus (luteus). Log phase cells of Micrococcus lysodeikticus (luteus) IFO 3333 autolyzed when incubated at 37 C in 0.01 M sodium-phosphate buffer pH 7.5. The enzyme involved in the autolysis was recovered mainly in an aqueous phase from cytoplasmic membranes and cytoplasmic materials treated with n-butanol, and proved to be an N-acetylmuramyl-L-alanine amidase. The autolysis of log phase cells suspended in autolyzing buffer was depressed by the addition of trypsin to the buffer.


A new method for extraction of extravasated dye in the skin and the influence of fasting stress on passive cutaneous anaphylaxis in guinea pigs and rats. A method for quantitative extraction of extravasated dye from the skin was studied in guinea pigs and rats. A simple method with a low cost and good recovery was established as follows; A piece of the skin containing extravasated dye was soaked overnight in a stoppered glass tube containing 1 ml of 1 N KOH at 37 C. Then, 9 ml of mixed solution of 0.6 N H3PO4 and acetone (5:13) was added to the tube. The tube was shaken vigorously for a few seconds and centrifuged at 3,000 rpm for 15 min. Absorbance of supernatant was measured at 620 nm. The recovery rate of the dye was about 95 per cent both in guinea pigs and rats. Using this method we observed that fasting stress significantly reduced the intensity of skin reaction induced by chemical mediators, heterologous PCA and especially homologous PCA in guinea pigs.


[Provocative gamma GT test in chronic pancreatitis? (author's transl)]. The increase of serum gamma-GT after hormonal stimulation has been reported as a new provocative test in the diagnosis of chronic pancreatitis. The test was suggested to be more sensitive than the serum amylase provocation test. In a clinical study of both tests in 83 consecutive patients the unsensitivity and the high rate of false positive results were demonstrated. Thus these provocation tests do not seem to be valuable in the diagnosis of chronic pancreatitis.


[Acetic acid, a catabolite repressor of cholinesterase synthesis by an Arthrobacter simplex culture]. Acetic acid was found to repress cholinesterase synthesis in the cells of Arthrobacter simplex var. cholinesterasus even at very low concentrations (0.1%). The repression is very stable. It is not eliminated by glucose or an organic acid of the Krebs cycle being added to the medium with acetic acid. The combination of acetic and butyric acids decreases the repression but does not eliminate it. The kinetics of cholinesterase synthesis was different in the cells grown on the medium with acetic acid and the cells cultivated on the medium with acetic acid and glucose, then washed and transferred to a fresh growth medium with glucose and acetylcholine as the sources of carbon.


[Disorder of porphyrin metabolism in thallium intoxication (author's transl)]. A 19-year old male ingested in suicidal attempt 750 mg of thallium. He developed the characteristic symptoms of thallium intoxication. During the acute phase the urinary excretion of porphyrins and porphyrin precursors was largely increased. The percentage distribution of the individual metabolites of heme synthesis revealed a preponderance of kopro- and uroporphyrin. This constellation (kopro- greater than uro- greater than tricarboxylic porphyrin) differs appreciably from that one in lead intoxication. The observation of increased urinary excretion of porphyrins and their precursors in a possibly particular spectrum in thallium intoxication is of special interest for differential-diagnostic reasoning. In each case of a toxic disorder of porphyrin metabolism thallium intoxication ought to be considered a possible cause.


[Effect of pH values of the medium unfavorable for growth on the physiological, morphological and cytological characteristics of a chemostat culture of Candida utilis]. The effect of growth inhibiting values of pH of the medium (pH 2.3 and 7.8) on physiological, morphological and cytological characteristics was studied with the chemostat culture of Candida utilis. Changes in the pH of the medium were accompanied with a decrease in the economic coefficient of the yeast cells, an increase in the respiratory quotient and the activity of phosphohydrolase. Phase-contrast micriscopy has shown that, at acid pH values, the cells were smaller than in the control (in the cells grown under optimal conditions) and rather uniform, oval or rounded, and contained large lipid granules. The presence of large lipid granules was confirmed by electron microscopy; vacuoles were smaller than in the control cells, but their number was greater than one; mitochondria stuck together and had one general outer membrane; the cell wall was 3--4 times thicker. At alkaline pH values of the medium, the whole cellular organization of C. utilis was disordered: the cells were larger, the size of vacuoles increased too much, and the structure became heterogeneous.


[Action of copper ions and of unfavorable medium pH values on protein and RNA synthesis by Candida utilis cells]. The effect of copper ions and unfavourable pH values of the medium on the incorporation of labelled precursors of protein and RNA was studied in Candida utilis. Specific inhibition of protein synthesis by copper ions and alkaline conditions was found. No specific inhibition of protein or RNA was detected at low pH values of the medium.


Enlarged acid-base and blood gas calculations by electronical data computing in the blood gas laboratory. A rapid anaysis of parameters of the acid-base equilibrium and blood gases during open heart surgery and emergency therapy is absolutely necessary. Computing of the several parameters of the acid-base status by slide rules or nomograms is time consuming and can be shortened by computer applications. The central blood gas laboratory consists of a blood gas analyzer for PO2, PCO2 and pH, an electronic desktop calculator, a four color X-Y-plotter and two data lines to the cardiac surgery unit and to the intensive care unit. The time needed for computing and feedback of the parameters could be decreased to one quarter. In addition to numerical data printout, a graphical representation of the several parameters is possible on a X-Y-plotter and includes the Rahn-Fenn-O2-CO2-Diagram with venous admixture, ventilation perfusion ratio, alveolar dead space ventilation and the standard and actual oxygen dissociation curve as well as the pH/HCO3- Acid-Base nomogram. Furthermore, a computer diagnosis of the actual disturbances can be plotted.


Characterisation of ribosomal proteins from HeLa and Krebs II mouse ascites tumor cells by different two-dimensional polyacrylamide gel electrophoresis techniques. Electrophoresis of ribosomal proteins according to Kaltschmidt and Wittmann, 1970a, b (pH 8.6/pH 4.5 urea system) yielded 29 proteins for the small subunits and 35 and 37 proteins for the large subunits of Krebs II ascites and HeLa ribosomes, respectively. Analysis of the proteins according to a modified technique by Mets and Bogorad (1974) (pH 4.5/pH 8.6 SDS system) revealed 28 and 29 proteins in the small subunits and 37 and 38 proteins in the large subunits of Krebs II ascites and HeLa ribosomes. The molecular weights of the individual proteins were determined by: 1. "three-dimensional" gel electrophoresis; 2. two-dimensional gel electrophoresis at pH 4.K/pH 8.6 in SDS. The molecular weights for 40S proteins ranged from 10,000 to 39,000 dalton (number average molecular weight: 21,000). The molecular weights for the 60S proteins ranged from 14,000 to 44,000 dalton (number average molecular weight: 23,000) using the "three-dimensional" technique. A molecular weight range from 10,000 to 38,000 dalton (number average molecular weight: 21,000) was obtained for the 40S subunits, whereas the molecular weights for the 60S ribosomal proteins (average molecular weight: 26,000) ranged from 12,000 to 69,000 dalton using the pH 4.5/pH 8.6 SDS system. The molecular weights Krebs II ascites and HeLa ribosomal proteins are compared with those obtained by other authors for different mammalian species.


Fidelity of conjugation in Saccharomyces cerevisiae. An efficient method for the production of synchronous zygotes in Saccharomyces cerevisiae is described. Cells were synchronised under defined conditions in either an a, alpha mixed culture or by incubation of each mating type in cell-free medium in which cells of the opposite mating type had been grown. Synchronised cells were allowed to fuse under defined conditions on filter membranes. This method was used to test the fidelity of conjugation in S. cerevisiae. Under conditions where cells of a or alpha mating type were in contact with up to 6 cells of each of two strains of opposite mating type, less than 1 multiple mating in 10(4) diploid matings occurred. It is concluded that is sexual conjugation in S. cerevisiae some process distinct from cell contact restricts cell fusion to paired combinations of conjugant cells.


Disturbance of the machinery for the gene expression by acidic pH in the repressible acid phosphatase system of Saccharomyces cerevisiae. When the pH of growth medium containing a limited amount of inorganic phosphate is kept below 3.0, cells of Saccharomyces cerevisiae produce repressible alkaline phosphatase but no repressible acid phosphatase. The same cells produce acid phosphatase immediately on shifting the medium pH to 4.0 or above. Like intact cells, spheroplasts prepared from cells grown at pH 3.0 or 4.5 in medium with a limited amount of inorganic phosphate in suspension begin production of acid phosphatase immediately after pH shift from below 3.0 to 4.0 whereas sheroplasts from cells grown in inorganic phosphate-rich medium showed a prolonged lag period (3 h). The enzyme formation on the pH shift was sensitive to cycloheximide. No significant differences could be detected in cellular growth or in incorporation of 3H-L-lysine or 14C-adenine between cells cultivated at pH 3.0 and 4.5. These results along with the fact that the expression of structural genes of repressible acid and alkaline phosphatases is controlled by a common genetic regulatory system, at least in part, indicate that the genetic regulatory system operates to express the structural genes even at low pH, though the expression of repressible acid phosphatase is interrupted. Coupled experiments of temperature and pH shifts with the temperature-sensitive mutants of the regulatory genes suggest that the acidic pH affects the function of the cytoplasmic products of those genes in the expression of the structural gene. Based on these observations, a revised model involving the simultaneous functioning of the regulatory factors was suggested for the genetic regulation of repressible acid phosphatase synthesis.


Differential mutagenicity of reaction products of various pyrazolones with nitrite. Four pyrazolones in frequent use, i.e. antipyrine (AP), aminopyrine (AMP), sulpyrine (SP) and isopropylantipyrine (IPA), were compared for their reactivity with nitrite and for the in vitro mutagenicity of their reaction products by Ames' reversion tests. In various acidic solutions at 37 degrees C, AP, AMP and SP were found to react easily with nitrite and yield various products including dimethylnitrosamine (DMNA) and 4-nitrosoantipyrine (4-NAP) in the cases of AMP and AP, respectively. When tested with Salmonella typhimurium TA100 and TA98 after lyophilization, the reaction products of AP (AP-N) were found to be mutagenic in both strains, while products of AMP and SP (AMP-N and SP-N) were mutagenic only in TA100. The presence of unknown ultimate mutagens, other than DMNA and 4-NAP, were evidenced in AP-N, AMP-N and SP-N. Incubation with S-9 mixture did not affect the mutagenicity of SP-N and decreased that of AP-N and AMP-N. In clear contrast to AP, AMP and SP, it was found that IPA remained essentially intact upon reaction with nitrite. No mutagenicity was detected with the reaction mixture (IPA-N) in either strain.


Substance P: binding to lipids in the brain. 1. Substance P (SP) could be extracted from brain homogenates with chloroform-methanol by a method which extracts all lipids. 2. SP could be transferred form this total lipid extract (TLE) into an aqueous solution at low pH values (2.0--3.0). 3. At higher pH values (5.5) SP could be transferred from an aqueous phase into an organic phase (chloroform:methanol, 2:1) and recombined with TLE (which was previously freed from endogenous SP) contained in this phase. The binding capacity of TLE for SP exceeded by far the amount of endogenous SP bound originally in the brain extracts. 4. Among the lipids present in TLE, phosphatidylserine was able to bind and release SP in a pH dependent manner. 5. It is suggested that SP bound to phosphatidylserine is the storage form of SP in the brain. The mechanisms by which it is released are still unknown. The possibility that the SP-receptor is also a phospholipid is considered.


The effect of some neuroleptics on the interaction of cortically and nigrally evoked potentials in the rat striatum. The interaction of cortico- and nigrofugal inputs to the striatum of the rat was investigated using the technique of evoked potentials. Repetitive, unilateral stimulation of the substantia nigra inhibited potentials which were evoked from the ipsilateral rostral cortex and recorded from the ipsilateral striatum. The inhibition was antagonized by low doses of various intraperitoneally administered neuroleptics such as: pimozide (0.1 - 0.2 mg/kg), haloperidol (0.1 - 0.5 mg/kg), chlorpromazine (0.5 - 2.0 mg/kg) and thioridazine (0.5 - 4.0 mg/kg). These findings are discussed in view of the existing controversy regarding the mode of action of dopamine in the striatum.


The treatment of anxiety with a polyfluorinated benzodiazepine derivative. Following 4 weeks of treatment with ORF-8063 a polyfluorinated benzodiazepine derivative, 8 hospitalized patients manifesting a primary pathology of anxiety showed marked general improvement. 2 other persons were treated, but for shorter periods: 9 and 14 days. Both are included in the pre-post analysis. Mean optimal dosage was 66.5 mg. The five instruments used to measure therapeutic effect showed pre- to posttreatment change with high level of statistical significance in serveral of the pathological factors. When measures of change are considered, patients showed more improvement related to psychic than somatic components of anxiety. Change data also indicates more patients improvement in anxiety than depression. Side effects reported more were dizziness, faintness and insomnia; these were reported in 8 patients. 6 patients noted drowsiness, and 4 noted excitement. 5 persons tolerated optimum dosages with no extreme reactions; 5 others (including the 2 subjects who terminated treatment early) were unable to maintain optimum dosages because of side effects.


Duration of apnea needed to confirm brain death. To determine the duration of respiratory arrest needed to attain a PaCO2 level high enough to provide maximal stimulation of respiration, we evaluated changes in PaCO2, PaO2 and apH during periods of apnea lasting as long as 10 minutes in 10 apparently brain-dead subjects. Before apnea, mean PaCO2 was 33 mm Hg. In seven subjects who did not breathe for 10 minutes, the mean rate of rise of PaCO2 was 3.2 mm Hg per minute. PaCO2 at 4 minutes was 50 mm Hg and at 10 minutes was 67 mm Hg. Three subjects breathed, two after less than 2 minutes of apnea, when PaCO2 was 47 and 54 mm Hg, and one after 4.5 minutes, when PaCO2 was 47 mm Hg. These data indicate: (1) that the threshold for respiratory stimulation may approach a PaCO2 of 60 mm Hg in patients with brain damage; (2) that the rate of increase in PaCO2 is such that, even in a normocapnic subject after 3 minutes of apnea, the PaCO2 may not be sufficiently high to stimulate respiration; and (3) if a patient is hypocapnic prior to the onset of apnea, PaCO2 may not reach 60 mm Hg even after 15 minutes. To confirm absolute apnea, then, blood gas monitoring is necessary for verification of normocapnia prior to the beginning of apnea. In the absence of blood gas determinations, no fixed period of apnea, sufficient in all cases to establish absolute apnea, can be ascertained.


Thiamine deficiency: selective impairment of the cerebellar serotonergic system. To explore the role of thiamine deficiency in synaptic transmission, the high-affinity uptake and release systems for putative neurotransmitters were studied in synaptosomal preparations isolated from the telencephalon, hypothalamus, and cerebellum of rats made thiamine deficient by diet or pyrithiamine. There was significant decrease in the uptake of serotonin by the synaptosomal preparations of the cerebellum. Although thiamine and its phosphorylated forms added in vitro did not restore the decreased serotonin uptake, the administration of the vitamin in vivo resulted in a significant reversibility of the inhibition of serotonin uptake, coinciding with dramatic clinical improvement. The study supports the possibility of an important serotonergic innervation of the cerebellum and suggests a selective involvement of this system in the pathogenesis of some of the neurologic manifestations of thiamine deficiency.