Bicarbonate ion transport: a mechanism for the acidification of urine in the turtle. The uricotelic turtle Pseudemys scripta acidifies the urine to a pH as low as 4 in the urinary bladder. Data in this report show that the mechanism of acidification in this bladder is the transport of bicarbonate ion from lumen to serosa, and that the temperature to which the turtles are adapted prior to the in vitro experiment largely determines the direction of the transmural carbon dioxide gradient observed. This temperature effect also serves to reconcile apparently disparate data that were previously reported. A new technique for the direct determination of the partial pressure of carbon dioxide was employed.


Oral cocaine: plasma concentrations and central effects. Cocaine (2.0 milligrams per kilogram) given by the oral route is at least as effective as the same dose given intranasally. Cocaine is not detected in the plasma until 30 minutes after oral administration, but peak plasma concentrations are similar after both routes. The subjective "highs" in man are greater after oral than after intranasal administration.


MEN type IIa syndrome: dilemmas in modern management. With the use of newer techniques, the multiple endocrine neoplasm II-A syndrome can now be diagnosed early in its evolution. This article discusses the clinical course of the syndrome, the diagnostic modalities employed, and the therapeutic interventions required. An approach to the individual patient with this syndrome is presented.


Muscle perfusion and the healing of below knee amputations. Muscle pH has been used to assess the perfusion of the muscles at the site of below knee amputation in 20 patients having ischemia at the periphery. The results show that a muscle pH above 7.0 did not correlate with the outcome of amputation but that, when the pH was below 7.00, healing was never achieved.


The role of fibrin formation in the pathogenesis of bacteremic shock in the primate. In this study, an attempt was made to elucidate further the role of intravascular fibrin formation in the pathogenesis of sepsis in the primate. It was found that injected live Escherichia coli caused death in primates within four to 11 hours as a result of microcirculatory failure and acidosis. Pretreatment with Arvin did not prolong the survival rate, probably because of an overloading of the reticuloendothelial system with fibrin degradation products. This study does not support an obligatory role for intravascular coagulation or fibrin formation in primate sepsis and coincides with an earlier report (6) from this laboratory on cats. Vascular damage and malfunction, secondary to mediators released by platelets, leukocytes, red cells or Hageman factor, are not ruled out.


Training and use of surgeon's assistants. "Physician extenders" of various types have been produced in the past few years. The University of Alabama in Birmingham has trained surgeon's assistants over the past 9 years, graduating 53 individuals after a 2-year course of didactic, laboratory, and clinical instruction. The graduates have performed satisfactorily, working with surgeons in the community setting and as part of highly specialized teams in large medical centers. One third work in general surgery, one third in cardiovascular thoracic surgery, and one third in other special areas. The authors conclude that these individuals will play an increasing role in allowing surgical therapy to be delivered in an efficient and effective manner.


Droperidol, its alpha-adrenergic blocking action on the aortic strip and inhibitory action on norepinephrine uptake of the adrenergic terminal of the left atrial strip of rabbit. In aortic strips, the dose-response curves for phenylephrine were obtained before and after addition of droperidol. Droperidol caused a parallel shift of the curves dose-dependently toward the right side and its grade of the shift was greater than caused by phentolamine at the same concentration. In the left atrial strip, the potentiation of contraction by tyramine was markedly depressed by droperidol. In the reserpinized preparation, the effect of tyramine was depressed markedly and there was no norepinephrine fluorescence. After incubation with norepinephrine they were restored to the same level as in non-reserpinized preparations. However, incubation with droperidol before norepinephrine blocked the restoration. It is postulated that droperidol may have both alpha-adrenergic blocking action and inhibitory action on norepinephrine uptake by adrenergic terminals.


The mechanism of urinary continence after prostatectomy. An anatomical and neuropharmacological study of the urethra was performed in 19 patients after prostatetectomy to investigate the mechanism of continence. Anatomically, the urethral segment responsible for the postprostatectomy continence was located distal to the open prostatic fossa in the external sphincter which is comprised of intrinsic striated muscle and is called arbitrarily rhabdosphincter by the author. This unique striated muscle in the rhabdosphinecter appears to be primarily under the alpha-adrenergic influence in view of the change in the urethral pressure profile observed following alpha-adrenergic drugs.


Pathology of germ cell tumors of the testis. Testicular germ cell tumors form a complex group of interrelated entities. Problems of nomenclature remain, but there does appear to be a close agreement between the major classifications in use. The identification of clinically useful markers for certain germ cell testicular tumors has provided a major impetus in this field. Information gained from the study of these markers has promoted an understanding of the histogenesis of germ cell tumors, has improved standard nomenclature, and has stimulated progress in patient care. Radioimmunoassay techniques are now frequently used to measure beta-human chorionic gonadotrophin and alpha-fetoprotein levels in evaluating the clinical course of patients with nonseminomatous germ cell tumors of the testis. It is hoped that the use of these markers will permit earlier detection of tumor recurrence, prompt treatment, and an improved survival rate. The knowledge of the production of alpha-fetoprotein by the human yolk sac has further refined tumor classification. It is against this general background that we have outlined the standard pathology of germ cell tumors of the testis.


Cytogenic studies of cryptorchid testes. Twenty-nine patients with unilateral or bilateral cryptochism underwent biopsies of the testis at the time of orchiopexy. Karyotype evaluation of fibroblasts obtained from tissue cultures of the biopsy specimen was completed counting a minimum of twenty-five cells. In 1 patient with multiple anomalies a 46,XYDq+ karyotype was identified. All other karyotypes were normal suggesting cryptorchism is not associated with abnormal testicular cytogenetics.


[Evaluation of metabolic profile tests using a computer]. A programme has been developed for the evaluation of metabolism profile tests in dairy cows by means of the TESLA-200 computer in the FORTRAN language. The computer evaluates and plots the metabolism profile from input data punched on tape in the IBM code by the Consul-253 electronic organization automatic device. The metabolism profile is represented as a column graph (histogram), the levels of individual parameters being confronted with the reference levels with a 95% confidence interval and the deviations are expressed in units defined as 1/2 of the confidence interval for each parameter. Calculation by this procedure enables the construction of a profile record in which any step beyond the 95% confidence interval in any parameters is seen very clearly. The computer also produces a table of exactly calculated values of deviations from the reference levels and makes it possible to determine the numbers of abnormalities for individual parameters in the animals tested.


[Intensity of acid-base reactions of the blood and cerebrospinal fluid in sheep after intraluminal stress induced by urea and acetic acid]. Acid-base indices of the blood and cerebro-spinal fluid were studied on Astrup apparatus (BMS 2) in an experiment with twelve, two- to three-year-old Merino sheep, being in the first half of the gravidity period, under clinical conditions after the administration of 0.8 g urea per 1 kg live weight followed by, in half an hour, the administration of acetic acid (8% vinegar, a dose of 2 ml per 1 kg live weight). Heparinized syringes were used to take blood samples anaerobically before the beginning of the experiment, then after 15, 30, 45, 60, 75, 90, 105, 120 minutes and after 4 and 24 hours. The fluid was taken by heparinized capillaries before the beginning of the experiment, then after 30, 60, 90 and 120 minutes and after 4 and 24 hours. The following indices were obtained, and/or calculated from nomographs: actual pH, carbon dioxide pressure--pCO2, base excess--BE, buffer bases--BB, standard and actual bicarbonate--SB, AB, total carbon dioxide--tCO2. After 15 minutes' interval since the administration of urea significant changes in the values of the indices, especially higher pH values and paradoxical pCO2 drop, were recorded. After the administration of acetic acid overcompensated acidosis of the organism occurred (accompanied even by mortality of the experimental sheep). Towards the end of the experiments the results illustrated basemia. A depressing effect of the lower pH on the toxicity of ammonia was also confirmed. The reaction of the fluid to the urea and acetic acid stress opposite to the venous blood was retarded and the values of the indices studied showed some variability.


Polypoid nonchromaffin paraganglioma of the duodenum. A polypoid tumor was surgically removed from the second part of the duodenum of a 56-year-old male. The main body consisted of large epithelioid cells arranged in an adenoma like pattern of strands and nests. These cells were argyrophil and had marked nonspecific esterase activity. Unmyelinated nerves with proliferated Schwann cells accompanied these epithelioid cells together with scattered gangliocyte like elements. Ultramicroscopically, the epithelioid cells were seen to contain round electron dense granules, 150 nm in diameter on average. The tumor is considered to be a nonchromaffin paraganglioma, as it probable developed from paraganglion cells associated with small arteries or branches of the vagus nerve, or from the undifferentiated pluripotent APUD cells of the duodenum.


[Artificial arterial hypotension during surgical treatment of intracranial aneurysms]. In studying the content of lactic and pyruvic acids in the cerebrospinal fluid of 31 patients during operation for intracranial aneurysms under halothane anesthesia, the authors established that the performance of these operations under protection of deep (40 mm Hg) and prolonged (up to 2 hrs 25 min) artificial arterial hypotension in an uncomplicated course of the surgical intervention was not attended with hypoxic damage of the brain. The danger of hypoxia developing became greater when acute disturbances of blood flow in the cerebral arteries (clipping of a vessel, spasm) occurred in the period of hypotension.


Dehydrogenation of indanol by rabbit liver 3-hydroxyhexobarbital dehydrogenase. 1. Among the several enzyme activities in rabbit liver cytosol able to dehydrogenate 1-indanol, only the main activity was not separable from 3-hydroxyhexobarbital dehydrogenase during purification including polyacrylamide gel disc electrophoresis. 2. Results of mixed substrate method indicated that the same enzyme catalyses the dehydrogenation of 1-indanol and 3-hydroxyhexobarbital. The ratio between the two dehydrogenation activities was almost constant as the enzyme underwent thermal inactivation. The Ki values of p-chloromercuribenzoate, the Km values for NAD+, and the Km values for NADP+ were very similar for the two dehydrogenations. These results lead to the conclusion that the same enzyme catalyses the dehydrogenation of 3-hydroxyhexobarbital and 1-indanol. 3. 1-Tetralol, 1-acenaphthenol, 9-fluorenol, thiochroman-4-ol and 4-chromanol also served as substrate of the enzyme, but 2-indanol, 2-tetralol, and trans- and cis-indan-1,2-diol were not oxidized. 4. Reversibility of the reaction was also confirmed using 1-indanone as substrate.


[Anabolic and catabolic enzymes of urea metabolism in a carbohydrate-utilizing strain of Candida guilliermondii]. The yeast "H" of the genus Candida guilliermondii can grow on hydrocarbons as the only source for carbon. Urea can serve as a nitrogen source for this yeast which lacks detectable urease activity. During urea metabolism ammonia has never been accumulated in the culture medium. However, transferring the yeast from complete urea-medium into an urea containing phophate-buffer, the degradation of urea continues and ammonia is accumulated as well as CO2 evolved. In cell-free extracts of the yeast urea amidolyase activity was detected in the presence of ATP, biotin and specific cations. Obviously, the synthesis of urea amidolyase is induced by urea and arginine and repressed by the catabolite ammonia. Similarly the synthesis of arginase is regulated by arginine and ammonia. The analytical data of the arginase action differ significantly in relation to the carbon source of the culture medium. Both the level of arginase and ornithine carbamyl-transferase change in a characteristic way during the batch-culture. From the lower level of arginase in relation to ornithine carbamyltransferase it can be concluded that especially in alkane-metabolizing yeast the arginine catabolism is not very intensive.


[On molecular biology of aging. 11th communication. Enzyme adaptation, age (author's transl)]. Age-dependent modifications of enzyme adaptations in response to a variety of environmental alterations are a biochemical expression of biological ageing. In this paper age-dependent changes of the adaptation of a number of enzymes are described. Experiments from our laboratory indicate that pyruvate kinase is more responsive to starvation and refeeding in young animals. The diminished response of pyruvate kinase in old age is caused by a decreased adaptability of the regulatory isoenzyme PK-L.


Effect of lime-pelleting on nodulation and yield of soybean grown in acid soil. An experiment for studying the effect of lime-pelleting of inoculated seed of soybean [Glycine max. (L.) Merr.] on nodulation, growth, and grain yield was undertaken in red sandy loam of Bangalore, having a pH of 4.0. The results indicated that inoculation alone increased significantly nodulation, plant dry weight, and grain yield. But inoculation plus lime-pelleting significantly increased the dry weight of nodules and plants, but not the grain yield. However, lime-pelleting was found to be beneficial though not essential in acid soils.


[Application of a TRICIN-buffered modification of the CMRL-1969 medium in cell and virus cultivation (author's transl)]. CMRL-1969 medium (Healy et al., 1971) was modified by using 0.02 molar N-[Tris-(hydroxymethyl)-methyl]-glycin (= TRICIN) instead of bicarbonate as the buffer substance. Several permanent cell lines and primary cell cultures did not show growth differences in the two medium variants. Like other non volatile buffers TRICIN abolishes the initial increase of the Ph in freshly split or newly fed closed vessel cultures, but in 0.02 M concentration maintains the same buffering capacity of the medium as compared to bicarbonate. Multiplication of Entero-, Adeno-, Herpes-, Influenza-A-, and Vaccinia-viruses was also not altered. The medium allows open vessel methods like microtiter- and plaque-tests to be conducted under normal atmosphere and seems especially useful when initial pH-values are to be set in the rank of 7.5 to 8.6. For microtiter neutralization tests with enteroviruses a medium adjusted to pH 7.6 and a cell density of 10(5) cells/cm2 was found optimal. Plaque formation of Coxsackie-B-viruses reached a maximal plaque-size and -number when the pH of the double concentrated medium was beyond 8.2.


Clotting and fibrinolytic activities of Bacteroides melaninogenicus. 106 strains of Bacteroides melaninogenicus ss. asaccharolyticus, intermedius and melaninogenicus were tested for production of extracellular blood clotting and fibrinolytic factors. 78.3% of tested strains caused clotting of rabbit plasma and 79.2% activated human plasminogen. Strains producing the clotting or fibrinolytic factor only were isolated. Both factors were quite active, as positive results for most strains were detected within one hour of incubation in controlled test system.


Studies on antituberculotic action of some phenothiazine derivatives in vitro. Five phenothiazine derivatives (chlorpromazine, levomepromazine, diethazine, promethazine and chlorpromazine) sulphoxyde were tested for antimycobacterial activity. The growth of Mycobacterium tuberculosis, M. bovis and M. butyricum was inhibited by chlorpromazine practically at identical concentrations. The minimum inhibitory concentrations for M. tuberculosis were: chlorpromazine and levomepromazine, 10 microgram/ml; diethazine and promethazine 20 microgram/ml, whilst chlorpromazine sulphoxyde was ineffective even at a concentration of 100 microgram/ml. Chlorpromazine and promethazine exerted a measurable bactericidal activity on M. tuberculosis at 50 microgram/ml; total destruction of the organism and loss of acid fastness in part of the cells were shown at 300 microgram/ml. Preliminary studies, in mouse experiments phenothiazine derivatives were ineffective.


The nature of clotting and fibrinolytic activities of Bacteroids melaninogenicus. From the supernatant of B. melaninogenicus ss. asaccharolyticus culture, a protein fraction was isolated by ethanol precipitation. The fraction was tested for the presence of clotting and fibrinolytic activities by application of quantitative techniques and specific substrates for measurement of prothrombin and plasminogen activation, and collagenase and elastase activity. It is postulated that ability of Bacteroides melaninogenicus ss. asaccharolyticus extracellular factors to clot fibrinogen and activate plasminogen, are due to a limited proteolysis by the proteolytic enzymes produced by this microorganism and not to the existence of specific B. melaninogenicus coagulase of plasminogen activator.


[Microbial growth on an expansion joint of a reservoir for drinkingwater (author's transl)]. In a clear water reservoir built in ready construction after a working-period of five months quite a lot of slime could be found on the expansion joint filled with tightening compound on the base of Thiokol. The covering slime could be removed easily with a rubber-screen. The microbiological analysis showed an extremely high number of colonies and additional the slime was secondarily settled with Protozoa. An influence on the water in the reservoir could not be found out under working conditions.


[On the determination of chlorine with a polarographic probe. 2nd communication: experimental arrangement and results under application (author's transl)]. The examination of the physical properties of the instrument showed several possibilities of meter-error mainly due to constructive disadvantages. After having found this a special experimental arrangement was worked out to exclude errors. Then good linear correlations of the meter-reading and the analytical DPD-Method for free Chlorine were found for pH-values ranging from 3.7 to 11. Further the results of a test under practical conditions in a swimming pool are shown and the possibility to discriminate different types of waters by their chlorine demand under constant-titration.


[Acid-base equilibrium in patients with tick-borne encephalitis and similar diseases]. The study is concerned with the acid-base balance in 92 patients with tick-borne encephalitis, tick-borne fever and meningitis of an obscure etiology. In many cases there was a shift in the direction of a metabolic acidosis, which was most vividly expressed in the meningitic form of tick-borne encephalitis. Some of the patients were treated by sodium bicarbonate which exerted a positive effect. The degree of changed indices of the acid-base balance and the time of their appearance helps in the diagnosis of tick-borne encephalitis correct evaluation of the severity of the condition and timely prescription of correction therapy.


[Use of neuroleptics for the treatment of chronic alcoholism (psychopathology of the attraction to alcohol)]. A psychopathological analysis of alcoholic drives in patients with alcoholism demonstrates that along with well-known negative symptoms of an alcoholic mental defect there is a productive psychopathological symptom in the clinical picture as well. Having this view, the author in 148 cases of alcoholism applied etoperasine (trilafon), and neuleptil (propericiasin)--neuroleptical drugs, possessing antipsychotic properties and relatively not toxic. As a result there was a distinct and rapid inhibition of pathological drives to alcohol with a good tolerance to the preparations. The use of these drugs activates the role of the patient in the process of treatment, increases the effectiveness of psychotherapy, and is technically simple in use. They may be prescribed for the prevention of relapses, arrest of bouts when other methods are ineffective.


Effect of extracellular pH and inhibitors on the gluconeogenesis and ammoniagenesis relationship in rat kidney cortex slices. 1. Gluconeogenesis from glutamine, fumarate, pyruvate, glutamine plus fumarate, and glutamine plus pyruvate, was generally higher at pH 7.1 than at pH 7.4 and 7.7, whereas ammoniagenesis did not depend on the pH of the medium. 2. The intermediates of the Krebs cycle decreased ammonia formation from glutamine, raising at the same time gluconeogenesis. 3. Arsenite, malonate, maleate, hydrazine and 2,4-dinitrophenol inhibited gluconeogenesis, and enhanced simultaneously ammonia formation irrespective of the pH of the medium.


Neuroleptic blockade of the effect of various neurotransmitter substances. The antagonistic effect of neuroleptics to acetylcholine, histamine, 5-HT, and noradrenaline was examined in various in vivo and in vitro models. Piflutixol, a new potent thioxanthene neuroleptic, markedly antagonizes the effect of dopamine, noradrenaline, 5-HT and to some extent histamine, whereas the affinity for muscarinic receptors was rather weak. Clozapine and chlorprothixene on the other hand, have high affinity for muscarinic receptors and also antagonize the effect of histamine and 5-HT, whereas clozapine was a weak antagonist of noradrenaline and dopamine when compared to the effect of piflutixol. Chlorprothixene, however, also exhibits a rather good antagonism of noradrenaline and dopamine. Haloperidol proved to be weak in all models when compared with the other neuroleptics examined. Flupenthixol specifically antagonizes dopamine and noradrenaline, whereas fluphenazine was a more potent antagonist of dopamine than of the other transmitters. The data show, that neuroleptic compounds possess very different profiles with regard to interaction with various neurotransmitter substances. It is suggested, that the rather potent anti 5-HT and antihistamine effects observed for certain substances may contribute to the central effect of these drugs.


Efficacy and side effects of flurazepam, fosazepam, and nitrazepam as sleeping aids in psychogeriatric patients. Efficacy and side effects of flurazepam 15 mg, fosazepam 60 mg, and nitrazepam 5 mg were studied in 17 psychogeriatric patients. The drugs were equipotent in maintaining sleep but nitrazepam had more side effects than the other hypnotics, and it induced a rebound insomnia after withdrawal. All hypnotics lost some of their efficacy towards the end of 7 days' administration. Patients with evident cerebrovascular disease were vulnerable to the side effects of the benzodiazepine hypnotics. The side effects did not correlate with the age of a patient. In addition, no correlations were found between the serum levels of fosazepam or its main metabolite and the side effects.


Three years' maintenance neuroleptic treatment in schizophrenia--before and beyond. A group of patients with schizophrenia, initially 67 patients, was studied over a period of 3 years. After three years 36 out of 67 patients were still on the same depot neuroleptic. The main aim was to describe and compare maintenance neuroleptic therapy using two depot neuroleptics, fluphenazine decanoate and pipotiazine palmitate, given monthly. Before the outpatient care the patients had participated in the department's comprehensive hospital treatment including depot neuroleptic medication. After a 1-year clinical trial with frequent assessments of the patients, significant symptom reductions were found on all rating scales. During the last 2 years of the study only drug therapy was given. Improvement concerning social function in the community and work level as well as the low-rated psychopathology noted at the start of study also persisted at the 3-year follow-up. The side effects were low in frequency and quality. These results show the clinical value of long-term maintenance treatment with depot neuroleptics. The results also confirm that the favourable effects of the hospital treatment demonstrated before the start of the clinical trial could be maintained. The possibilities of further improving aftercare and outpatient treatment beyond medication alone are discussed.


How schizophrenic patients change during 3 years' treatment with depot neuroleptics. A group of patients, initially 67 individuals, with chronic schizophrenia were studied on repeated occasions during 1 year and followed up after 3 years. The patients were given depot neuroleptics, either fluphenazine decanoate or pipotiazine palmitate, at intervals of 1 month. The symptom scores from three rating scales were subjected to factor analysis. Four factors were found to explain the variance satisfactorily: one comprising psychopathological symptoms specific for schizophrenia, one relating to contact disturbances, one psychomotor activity and one representing neurotic symptoms. Analysis of these factors revealed certain differences between the treatment groups over time and demonstrated the effect of combination of psychotherapy and neuroleptic drugs in a subgroup of patients. This type of analysis of treatment results might contribute to improving our knowledge of rehabilitation of schizophrenic patients and help us to draw up giudelines for selection of suitable measures.


Auricular tyrosine hydroxylase and dopamine-beta-hydroxylase activities and noradrenaline content in ischaemic heart disease. Assessments were made of the tyrosine hydroxylase (TH) and dopamine-beta-hydroxylase (DBH) activities as well as the noradrenaline (NA) content of samples excised from right auricular tissue during cardiac surgery on a total of 55 patients with ischaemic heart disease (IHD), valvular heart disease (VHD), uncomplicated atrial septal defect (ASD) or congestive heart failure (CHF). The NA content was significantly higher in the IHD group than in the other three groups. The TH activity was highest in the IHD group although the difference was statistically significant only compared with the ASD and CHF groups. The DBH activity was also highest in the IHD group, but again the difference was statistically significant only compared with the ASD and CHF groups. In the whole material there was a significant positive correlation between the NA content and TH or DBH activity, as well as between TH and DBH activity. In the IHD group there was a significant positive correlation between heart volume and TH activity. The results suggest that at least compared with ASD and CHF, the sympathetic tone is relatively high in IHD, possibly involving an enhanced NA turnover.


Cyclic nucleotide metabolism in solid tumor tissues. The examination of the regulation of the system of 3'-5' cyclic nucleotide monophosphates has only begun in cancer tissues. In human cancers, these studies are notably non-existent. However, in animal cancers, especially the Morris hepatomas, enough data has been gathered that, while risky, certain trends seem to begin to appear. Cyclic AMP is constant or lowered, while cyclic GMP is elevated in the fast growing hepatomas. Regulation of adenylate cyclase by protein hormones is reduced, while regulation by epinephrine may be increased. Binding of glucagon is decreased in the fast growing hepatomas. Guanylate cyclase, while being predominantly cytoplasmic in the normal liver, is predominantly membrane bound in the tumors. The liver enzyme is also readily stimulated by several chemical carcinogens. The cyclic GMP phosphodiesterases are decreased in these tumors; while the cAMP phosphodiesterases are increased. Although the cyclic nucleotide dependent protein kinases (histone as substrate) are altered in the hepatomas, observations of unique cyclic nucleotide binding proteins or cAMP independent protein kinases in cancer tissues may be of even greater significance for the development of or the maintenance of the neoplastic state of cells.


Quantitative effects of salycylhydroxamic acid and glycerol on Trypanosoma brucei glycolysis in vitro and in vivo. During anaerobic glycolysis in vitro in the presence of salicylhydroxamic acid, Trypanosoma brucei brucei converts glucose to equimolar amounts of glycerol and pyruvate as end products. Glycerol, whether generated endogenously and pyruvate as end products. Glycerol, whether generated endogenously or added exogenously, can inhibit anaerobic glycolysis sufficiently in vitro to result in cell death. The concomitant administration of salicylhydroxamic acid and glycerol to rats infected with T. brucei brucei results in a rapid clearance of parasitemia. Our results clearly demonstrate a new and approachable chemotherapeutic target for African trypanosomes.


Effect of beta-lapachone on hydrogen peroxide production in Trypanosoma cruzi. beta-Lapachone, an antimicrobial agent, markedly increase the generation of H2O2 in intact Trypanosoma cruzi epimastigotes (Y strain). Increase in H2O2 was determined by the horseradish peroxidase-H2O2 Compound II formation as well as by a cytochemical technique.


[Data on the life cycle of Ribeiroia marini guadeloupensis n. ssp., a trematode sterilizing Biomphalaria glabrata in Guadeloupe]. Data on the life cycle of Ribeiroia marini guadeloupensis n.ssp., Trematode sterilizing Biomphalaria glabrata in Guadeloupe. Maintenance of the life cycle with a view to an eventual control of the populations of Molluscs. In Guadeloupe (French West Indies), the definitive hosts of Ribeiroia marini guadeloupensis n.ssp. are the two species of rats: Rattus rattus and R. norvegicus. The first intermediate host is Biomphalaria glabrata, the vector of intestinal schistosomiasis. The second intermediate host is a fish (Tilapia mossambica, Poecilia reticulata). The different stages of the life cycle are described; the transmission is studied experimentally and in natural conditions. A method for producing eggs of this Trematode in large quantities is described.


Alteration of amicrofilaremia in Dipetalonema viteae infected hamsters with immunosuppressive drugs. Dipetalonema viteae-infected hamsters with amicrofilaremic infections were subjected to immunosuppressive therapy. Methyl prednisolone acetate caused the most severe recrudescence of microfilariae while cyclophosphamide caused a low level, transient microfilaremia. Saline injected control hamsters remained amicrofilaremic. Neither drug influenced the number of adult worms recovered at necropsy in the treated hamsters compared with control hamsters.


Serodiagnosis of human cysticercosis by microplate enzyme-linked immunospecific assay (ELISA). The micro-ELISA described in this report is simple and can give objective results for the serodiagnosis of human cysticercosis. The visual assessment of results was satisfactory. The ELISA results showed a remarkably good correlation with IHA. It gave good reproudcibility. Cross-reactions were eliminated by using whole worm antigen instead of the cysticercus.


Responses of Anopheles minimus to DDT residual spraying in a cleared forested foothill area in central Thailand. Anopheles balabacensis balabacensis and Anopheles minimus are the main malaria vectors in Thailand. In a cleared forested foothill area in the central part of the country A. minimus was the most prevalent anopheline species found, only 6 specimens of A. b. balabacensis being collected over a 3-year period. Cattle were scarce in the area, tractors being largely used for working in the fields. This situation contributed to high man-vector contact. A minimus occurred throughout the year, with a major peak of density in the dry cool season and a smaller peak in the wet season. The contact of A. minimus with man was much higher outdoors than indoors, and studies showed the species to be an early biter, especially in the dry season, thus increasing the chance of man-vector contact. DDT spraying appeared to reduce considerably the estimated vectorial capacities, however, this effect was not maintained and malaria transmission was not interrupted. Trials with supplementary or alternative attack measures are therefore indicated in this particular ecological situation.


[Studies on an isolated tribe in the central highlands of Irian-Jaya (Indonesian New Guinea) within the field of tropical medicine. I. Clinical studies]. Reports on medical research examinations in the field of tropical medicine of 130 indigenous, who were examined in the central Highland of West-Irian during an interdisciplinare research project. The inhabitants of this markedly isolated area are pygmies of a strong, well proportioned build of the body and in a relative good general condition. No avitaminoses, nor symptoms of malnutrition, nor tuberculosis were discovered, in spite of an only moderate intake of albumine and carbohydrates. All examined indigenous show a distinct diastase of the musculi recti, 51.55% an enlargement of both glandulae parotes, 47% an enlargement of the lymph nodes in the differemt regions, and 11.52% an enlargement of the liver. Distinct was a violent endemy with intestinal worms and some extreme anaemia. A later publication will deal with the extensive laboratory examinations which are partly not yet completed.


Radioimmunoassay of influenza A virus haemagglutinin. I. Preparation and properties of radioactive 125I-labelled bromelain-released haemagglutinin. Haemagglutinin released from influenza A virus recombinant MRC11 [antigenically identical to the strain A/Port Chalmers/1/73 (H3N2)] by bromelain treatment and purified by rate zonal centrifugation (further on B-HA) was examined for eventual contamination by neuraminidase. According to specific enzymatic activities corresponding to MRC11 virus and B-HA alone respectively, B-HA contained less than 0.1% of enzymatically active neuraminidase orginally present in the virus. Gel double diffusion tests, specifities of rabbit antisera induced by B-HA, as well as radioimmunoprecipitation experiments demonstrated that B-HA was devoid of any antigenically active neuraminidase. Precipitation of 125I-labelled B-HA with antisera to influenza virus recombinants with N2 neuraminidase has been evidently caused by antibodies to host antigenic determinaant(s) present in these sera. With respect to purity as well as radioimmunoprecipitation properties, B-HA is quite suitable for radioimmunoassay experiments.


Effect of 5-bromo-2'-deoxyuridine on the in vitro DNA synthesizing activity of polyoma virus-infected cells. The in vitro synthesizing activity of uninfected and polyoma virus-infected mouse embryo (ME) cells was investigated by measuring the incorporation of 3H-thymidine (dTR) into acidinsoluble material in cell-free extracts. The character of the enzyme reaction was described in uninfected cells and an activating effect of 0.01 M dithiothreitol (DTT) was observed. A variation of the enzyme activity in the course of cultivation of ME cells in primary culture was demonstrated. The induction of the DNA synthesizing activity due to polyoma virus infection was markedly increased in the presence of 5-bromo-2-deoxyuridine (BUDR), what emphasizes the importance of the de novo synthesized viral and/or cellular DNA in the regulation of induction enzymes participating in DNA synthesis.


Some properties of the adenosine triphosphatase associated with herpes simplex virus and nuclear membrane of host cells. An enzyme capable to split adenosine triphosphate (ATP) was shown to be firmly associated with mature herpes simplex virus particles purified from infected rabbit lung (ZP) cells. The enzyme localized in the viral envelope was markedly activated by bivalent cations, to the largest degree by Mg2+ at a pH optimum of 7.8--8.0. Na+ and K+ ions neither separately nor together showed any activating effect. Enzyme activity was not sensitive to the action of ouabain. No adenosine diphosphatase (ADPase) and adenosine monophosphatase (AMPase) activities were observed. ATPase activity was competitively inhibited by ADP. AMP and inorganic phosphate were without effect. The ATPase of nuclear membranes isolated from ZP cells exhibited similar properties but behaved differently to the action of sodium dithionite, dinitrophenol, oligomycin and gramicidin, as well as on heat inactivation. The origin of the virus enzyme is discussed.


Dengue viral antigens in host cell membranes. Membranes from type 2 dengue virus (DEN-2) infected BHK cells, when used as immunogen, elicit antibodies detectable by complement fixation, hemagglutination inhibition and plaque reduction neutralization tests with suckling-mouse-brain-derived DEN-antigens. Membrane fractions from infected cells separated by sucrose density gradient centrifugation were employed as antigen in serological tests with anti-DEN-2 infected suckling mouse brain hyperimmune mouse serum and hyperimmune mouse ascitic fluid. Antigen (s) capable of fixing complement were associated with all membrane fractions, including the plasma membrane, whereas antigen capable of reacting with antihemagglutinin was largely confined to the two membrane bands consisting of (1) a mixture of rough and smooth endoplasmic reticulum (2) predominantly rough endoplasmic reticulum.


Studies of cell-mediated immunity to Junín virus. Junín virus is uniformly lethal in newborn mice, but fails to kill adult animals. But there is direct evidence that in older mice the immunological mechanism occurs when certain conditions are fulfilled. The present results showed that in Junín virus infection of mice, the development of the immunological mechanism occurs irrespectively of age. The induction of a regular cell-mediated immunity appears to be intimately related to multiple injections of the virus. These results were confirmed by the 51Cr release assay and by the effect of the transplanted sensitized cells. The extent of infection in the brain is another variable to be considered. Only when the virus has reached high titres and has been concentrated in the brain, the damage done by the immune attack is lethal. On the other hand the data obtained suggest that Junín virus-sensitized spleen cells do not possess the ability to transfer significant antiviral effects to recipient preinfected newborn mice.


Antiviral pyrimidine derivatives: binding with biopolymers and photosensitization. Two antiviral pyrimidine derivatives, known to partially inhibit RNA dependent RNA polymerase of Mengo virus in a cell-free system, were found to bind with biopolymers. The antiviral compounds, at concentration of 100 and 50 muM and lower, showed complete inhibition of the virus-induced cytopathic effect and plaque reduction. Both compounds showed fluorescence emission with a maximum at about 500 nm under the influence of UV light of the wavelength of 366 nm. The fluorescence intensity increased upon addition of aqueous solutions of DNA, RNA and human serum albumin. This indicates that both compounds are capable of binding with nucleic acids and proteins. Based on the binding experiments alone it cannot be decided whether interference with nucleic acid template activity, enzyme function, or both are responsible for the virostatic action. Irradiation with violet light considerably enhances the virostatic activity, a fact that may be caused by photodynamic processes.


Effect of 3-[bis-(2-hydroxyethyl)-amino]-acetophenone-[4.5-diphenyl-oxazolyl-(2)]-hydrazone (IMET 98/69) on lethal mengo virus encephalitis. 3-[Bis-(2-hydroxyethyl)-amino]-acetophenone-[4.5-diphenyl-oxazolyl-(2)]-hydrazone (IMET 98/69) was found to afford maximum "rates of protection" in intraperitoneally (i.p.) or intranasally (i.n.) induced Mengo virus encephalitis in mice when administered once daily at doses of 1 mmole/kg (456.5 mg/kg) subcutaneously (s.c.) or 4 mmoles/kg (1 826 mg/kg) perorally (p.o.). Three days of treatment were sufficient if started on the day before or at the time of virus inoculation. Initiation of treatment 6 hours after inoculation was no longer effective. In a remarkable number of brains from infected and treated mice no virus was detectable, while in the remaining brains the appearance of virus was strongly delayed, and its amount significantly reduced.


Reproducible plaquing system for rhinovirus serotypes in HeLa cells--agarose suspension. HeLa cell--agarose suspension method provides an excellent plaquing system for assay of rhinovirus serotypes which ordinarily produce indistinct plaques on cell monolayers. The variable plaque size of cloned virus progeny does not involve antigenic change in phenotype as shown by plaque neutralization test. The system has proved useful for accurate and reproducible plaque assay of rhinoviruses, assay of serum neutralizing antibodies and cloning of viruses. The plaques produced are large, clear and reproducible. The factors which influence the plaquing of rhinoviruses in this system are the age of the cells, cell density and pH of the medium.


Use of gel precipitin test in the diagnosis of cytomegalovirus infections. Cytomegalovirus (CMV) antibody was detected in human sera by the gel precipitin test. The results were closely related to those obtained by complement fixation. The simplicity of the test makes it useful for the routine serological diagnosis of CMV infection.


Preparation of rubella virus haemagglutinating antigen by polyethlene glycol precipitation. Polyethylene glycol (PEG) was used for precipitation of rubella virus haemagglutinating (HA) antigen from the fluid phase of infected Vero cell cultures, maintained with or without calf serum. In both cases potent antigen preparation were obtained. When the antigen was precipitated from serum-containing medium, higher titres were obtained, provided that substrate or final product were treated with Tween 80 and ether. HA antigen obtained by PET precipitation was entirely valuable in the haemagglutination-inhibition test, as compared with antigens prepared by other methods.


Isolation of Tettnang virus from Ixodes ricinus ticks in Czechoslovakia. Three virus strains closely related to Tettnang virus were isolated from nymphs and males of Ixodes ricinus tick in Moravia and Slovakia. The virus isolates were lethal for suckling mice on intracerebral (i.c.) inoculation but the virus titres in the brain were low. The incubation period of 5--12 days in the first mouse passage was shortened to 3 days in further passages. Infection of mice was accompanied by ultrastructural changes in the cytoplasm of neurons. Groups of spherical virus particles 75 to 80 nm in diameter occurred within simple vacuoles.


The nature of the renal response to chronic disorders of acid-base equilibrium. The rate of acid excretion by the kidney appears to be determined by factors regulating the site and the rate of sodium reabsorption, rather than by a homeostatic mechanism that responds to systemic pH. This hypothesis, although unconventional, is supported by much experimental evidence, and it accounts for a wide variety of clinical and physiologic findings that heretofore have been difficult or impossible to explain.


Sympathetic responsiveness and antihypertensive effect of beta-receptor blockade in essential hypertension. The relationship between sympathetic responsiveness and the blood pressure reduction induced by long-term beta-blockade was assessed in patients with essential hypertension. The increase in plasma noradrenaline concentration during physical exercise was used as an index of sympathetic responsiveness. The cardioselective beta-blocker, atenolol, was given to 16 patients with sustained benign essential hypertension for five weeks at a dose of 200 mg/day. Atenolol induced a marked decrease in blood pressure and pulse rate during recumbency, orthostasis and exercise concomitant with a marked increase in plasma noradrenaline concentration (p less than 0.0125) and a pronounced decrease in plasma renin concentration (p less than 0.01). The ratio of plasma noradrenaline during exercise to the base line concentration correlated significantly with the subsequent decrease in mean arterial blood pressure induced by beta-blockade (r = 0.840; p less than 0.001). A less significant correlation was observed between the plasma renin concentration and the subsequent decrease in mean arterial pressure (r = 0.542; p less than 0.05). The results obtained indicate that sympathetic responsiveness is an important determinant of blood pressure response to beta-blockade induced by atenolol.


Sodium wasting, acidosis and hyperkalemia induced by methicillin interstitial nephritis. Evidence for selective distal tubular dysfunction. A 61 year old male patient was studied who manifested dehydration, azotemia, acidosis and hyperkalemia six weeks after exposure to methicillin. Thyroid and adrenal glucocorticoid and mineralocorticoid function were normal. The dehydration was found to be caused by a profound sodium-losing nephropathy; urinary sodium ranged from 78 to 101 meq/day during a salt restricted diet. A distal renal tubular acidosis and a quantitively impaired ability to excrete potassium were also found. These defects were relatively unresponsive to mineralocorticoid or prednisone therapy. A renal biopsy specimen showed an interstitial nephritis which selectively affected distal tubules and was thought to be secondary to methicillin. The data suggest functional impairment specific for the distal tubule, but with only a modest decrease in the glomerular filtration rate.


New concepts in dysmenorrhea.

New concepts in dysmenorrhea. The etiology of primary dysmenorrhea, which is the most common gynecologic complaint and cause of lost working hours, remains obscure but merits careful scientific investigation. Recent studies suggest that increased endometrial prostaglandin production and release may be responsible for dysmenorrhea. Prostaglandins cause myometrial contractility that, if excessive, leads to uterine ischemia and pain. This hypothesis has led to clinical trials of antiprostaglandin agents such as indomethacin and fenamates, which inhibit the synthesis of prostaglandin through the prostaglandin synthetase system as well as antagonize their action at the cell receptor level. The good response of dysmenorrhea to other conventional forms of therapy such as oral contraceptives and dilatation of the cervix can be partly explained on the basis of a reduced level of prostaglandins in the menstrual fluid with such therapy. There is a definite need for further evaluation of the antiprostaglandin compounds in the treatment of dysmenorrhea so that sound formulations can be evolved for the elimination of this incapacitating disorder.


Subtle sequelae of brain damage. Perplexity, distractibility, and fatigue. Subtle problems of perplexity, distractibility, and fatigue accompany all kinds of brain injury for they appear to result from disruption of accustomed neural pathways and loss or change of mental function. Emotional disturbances may mask these subtle problems, but they can also result from them. Clinical experience indicates that these problems are more apt to become stressful when the patient misinterprets or copes ineffectively with them. That these common problems of brain injured adults may be overlooked in the usual clinical examination was shown in a comparison of clinical records of 50 patients referred for neuropsychological consultation with records of 46 patient-participants in a longitudinal neuropsychological study. Significantly more reports of these problems appeared in the latter group of records. However, consultation records did reflect emotional distress. Irritability, depression, or anxiety affected all but six consultation patients and appeared with equal frequency among working patients as among those unable to work or needing full-time care. This suggests that some of their emotional distress resulted from less obvious problems than those impairing mobility, strength, or competency. Counseling can reduce the patient's vulnerability to the psychologically crippling effects of perplexity, distractibility, and fatigue. Specific recommendations for patient and family counseling are offered.


The syndrome of post-splenectomy fulminant sepsis. Case report and review of the literature. Three years after splenectomy, a middle-aged woman had two separate episodes of fulminant bacterial sepsis. She recovered each time with prompt and appropriate treatment. Her immunologic system was examined and found to be normal with respect to antibody formation against diphtheria and tetanus toxoid, granulocyte killing of staphylococci and serum opsonizing activity. Granulocytes, bursa-equivalent (B) and thymus-dependent (T) lymphocytes and serum immunoglobulins were quantitatively normal. She produced antibody against subcutaneously injected polyvalent pneumococcal vaccine. She demonstrated cutaneous anergy. The literature on this syndrome was reviewed in an attempt to ascertain why hyposplenic patients are subject of fulminant bacterial, chiefly pneumococcal, sepsis. The probably explanation is the delayed production of antibodies against the phagocytic-resistant capsule of certain bacteria which the host had not previously encountered. The incidence of this syndrome appears to be on the order of 0.5 to 1.0%/year for splenectomized older children and adults. As the syndrome is seen chiefly in splenectomized or othewise hyposplenic patients, a causal relationship seems to exist.


Smoking education programs 1960-1976.

Smoking education programs 1960-1976. This paper is a review of published reports, in English, of educational programs designed to change smoking behavior. Attempts to change the smoking behavior of young people have included anti-smoking campaigns, youth-to-youth programs, and a variety of message themes and teaching methods. Instruction has been presented both by teachers who were committed or persuasive and by teachers who were neutral or presented both sides of the issue. Didactic teaching, group discussion, individual study, peer instruction, and mass media have been employed. Health effects of smoking, both short- and long-term effects, have been emphasized. Most methods used with youth have shown little success. Studies of other methods have produced contradictory results. Educational programs for adults have included large scale anti-smoking campaigns, smoking cessation clinics, and a variety of more specific withdrawal methods. These methods have included individual counseling, emotional role playing, aversive conditioning, desensitization, and specific techniques to reduce the likelihood that smoking will occur in situations previously associated with smoking. Some of these techniques have produced poor results while studies of other methods have shown inconsistent results. The two methods showing the most promise are individual counseling and smoking withdrawal clinics.


Neuroendocrine carcinomas of the colon. Ultrastructural and biochemical evidence of their secretory function. Four cases of malignant colonic tumors diagnosed by light microscopy as "small cell undifferentiated carcinomas" were shown by electron microscopy to have neurosecretory-type granules. Biochemical analysis of tumor tissue extracts disclosed the presence of considerable levels of VMA and catecholamines in all tumors; 5-HIAA was present in one tumor. Clinically, there had been no signs or symptoms attributable to those or related substances. Similar observations have been reported in a variety of neuroendocrine neoplasms; for example, the demonstration of neurosecretory-type granules and determination of amine or peptide materials in tumor tissue or body fluids may not be necessarily reflected in clinical hormonal syndromes or obvious metabolic abnormalities. Our structural and biochemical observations indicate that, regardless of clinically evident hormonal activity or lack thereof, some small cell "undifferentiated" colonic cancers derive from APUD elements, and therefore they should be classified within the group of neuroendocrine carcinomas. The evident secretory capabilities of these carcinomas suggest obvious diagnostic possibilities and could conceivably lead to a reappraisal of current therapy.


Biochemical tests for the detection of alcoholism: comparison of plasma alpha-amino-n-butyric acid with other available tests. The measurement of AANB relative to leucine was found to be a useful biochemical test for the detection of chronic alcohol consumption. This test, in combination with GGTP determinations, provided a sensitive means of detection (81% positive) for chronic alcohol consumption with a minimum of false positive determinations (2%).


Pain and clinical thrombophlebitis following intravenous diazepam and lorazepam. Eighty-seven per cent of surgical patients receiving undiluted diazepam experienced pain on injection while 6-16%, depending on the dose, manifested evidence of clinical thrombophlebitis. This was improved when diazepam, 10 mg, was diluted to 20-40 ml with intravenous solution. In contrast, lorazepam appeared to have minimal irritative or injurious effects on veins whether undiluted or diluted. In view of these results and clinical studies reporting a higher patient acceptance of lorazepam than diazepam, lorazepam may be a superior drug for use in anaesthesia.


Arterialized capillary blood used to determine the acid-base and blood gas status of dogs. Blood samples were collected by microhematocrit tube from posterior medial margin of the shaved, but otherwise untreated, canine ear. Acid-base and blood gas values of these samples were compared with the values of samples obtained simultaneously from the carotid artery. The arterialized nature of capillary blood from the canine ear was demonstrated under various degrees of chemical restraint and during conditions of extreme hypoxic acidosis to hyperventilatory alkalosis. Once a week determinations of acid-base and blood gas status with such arterialized capillary blood from a group of awake dogs showed within-subject variance to be significantly less (P less than 0.05) than between-subject variance; thus, uniqueness of individual dogs was reliably revealed. This technique also was used to demonstrated breed differences for acid-base and blood gas characteristics.


Type-specific pneumococcal respiratory disease in the elderly and chronically III. Type-specific pneumococcal respiratory disease was studied in a chronic-disease hospital during a 27-month period. Isolates from 50 patients with pneumonia and from 24 patients with chronic bronchitis were available for typing. Vaccine types were isolated from 74 per cent of patients with pneumonia and from 42 per cent of patients with chronic bronchitis. Pneumococcal types isolated from 5 of 8 patients with bacteremia and from 6 of 9 patients who died were also included in the vaccine. The data suggest that, theoretically, closed populations of elderly and chronically ill patients would benefit from vaccination in an attempt to control pneumococcal pneumonia. Less clear is the potential role vaccination in patients with chronic bronchitis.


Subacute thyroiditis with increased serum alkaline phosphatase. Three patients had subacute thyroiditis and elevated serum alkaline phosphatase, presumably related to the thyroiditis. Concomitant elevation of the serum gamma-glutamyl transferase suggested that the alkaline phosphatase was of hepatic origin. The elevation of the serum alkaline phosphatase could not be definitely related to the degree or duration of elevation of the serum thyroxine. The combination of elevated serum alkaline phosphatase with the systemic symptoms of subacute thyroiditis may obscure the diagnosis, especially if patients have little or no neck pain.


Clinical and biochemical heterogeneity of depressive disorders. Indirect evidence, mostly pharmacologic, has suggested a role for brain neurotransmitter amines such as norepinephrine in the production of depression or mania. Clinical investigations have supported this concept but also indicate that depression is probably a biochemically heterogeneous group of illnesses. There may be a clinically, biochemically, and pharmacologically definable subtype of depression in which there is a disorder of norepinephrine metabolism or disposition in brain. I review here the experimental data from which this hypothesis is derived.


Relationship of pH of blood and aqueous with vitamin C. Blood pH, aqueous pH, and intraocular tension were noted in 20 albino rabbits before and after administration of vitamin C in oral dosage of 7.5 gm/kg/day (in 4 equal divided doses for one week). A significant difference was noted in changes of blood pH and fall in intraocular tension but aqueous pH did not show any change. In 20 glaucoma patients there was no significant change in pH of blood of glaucoma after administration of vitamin C in 3.5 gm/kg body weight per day by oral route for one week (in 4 equal divided doses).


Origin of an elevated plasma alkaline phosphatase activity in non-jaundiced patients. Samples from 260 non-jaundiced patients with elevated plasma alkaline phosphatase activities were analysed for gamma-glutamyltransferase and 5'-nucleotidase activity, and for alkaline phosphatase isoenzyme pattern. The plasma gamma-glutamyltransferase activity was found to be a more sensitive index than that of plasma 5'-nucleotidase in confirming the presence of a liver component of the elevated plasma alkaline phosphatase. If the gamma-glutamyltransferase level is normal it is probable that the increase in plasma alkaline phosphatase activity is of bone origin. However, and elevated gamma-glutamyltransferase result does not exclude a bone component; in this situation plasma alkaline phosphatase isoenzymes should be estimated. The causes of elevated activities of plasma alkaline phosphatase, 5'-nucleotidase and gamma-glutamyltransferase, found in this investigation were generally the same as those found by other worker. The effect of treatment by drugs on gamma-glutamyltransferase, an inducible enzyme, needs more investigation.


Purification and some properties of riboflavin synthetase from Bacillus stearothermophilus ATCC 8005. A riboflavin synthetase was purified 51-fold from a thermophilic organism, Bacillus stearothermophilus ATCC 8005, that grew at 40 to 72 degrees C. Some of the properties of the enzyme are: (i) its temperature optimum was 95 degrees C, and the activity was negligible below 40 degrees C; (ii) the Arrhenius plot of the initial reaction rates was concave upward, with a break at 65 degrees C, and the apparent activation energies below and above 65 degrees C were 4.2 X 10(4) and 6.7 X 10(4) J/mol, respectively; (iii) the enzyme was fairly stable up to 60 degrees C without 6,7-dimethyl-8-ribityllumazine; this substance protected the enzyme from inactivation above 60 to 97 degrees C; (iv) the pH range for stability was 6.0 to 10.0 at 26 degrees C and 6.3 to 7.6 at 55 degrees C; (v) the enzyme was highly resistant at 26 degrees C to denaturation in 8 M urea, but the tolerance was extremely low at 55 degrees C; (vi) its molecular weight was estimated at 45,000; (vii) the Km for 6,7-dimethyl-8-ribityllumazine was 23 micrometer at 55 degrees C and 29 micrometer at 75 degrees C; (viii) its pH optimum was 6.7 to 7.2; (ix) 6-methyl-7-hydroxy-8-ribityllumazine was a competitive inhibitor (Ki = 0.18 micrometer); (x) the activity was sensitive to heavy-metal ions and thiol reagents; (xi) the enzyme did not require cofactor or a carbon donor; and (xii) the molar ratio of 6,7-dimethyl-8-ribityllumazine consumption to riboflavin formation was 2 throughout the entire reaction. Properties i through vi distinguish this enzyme from riboflavin synthetases purified by other investigators from mesophilic organisms, Ashbya gossypii, Eremothecium ashbyii, Escherichia coli, yeast, and spinach.


A supplement to alkaline phosphatase fractionations: utilization of gamma-glutamyl transpeptidase and hydroxyproline assays. Fractionation of serum alkaline phosphatase by either biochemical or electrophoretic techniques is often insufficient to determine the source of the elevated serum enzyme. In 31 unselected patients with high serum alkaline phosphatase a simultaneous determination of serum gamma glutamyl transpeptidase and urinary hydroxyproline excretion rates was performed, in addition to urea denaturation and L-phenylalanine inhibition tests. Of 25 patients with definite diagnosis, the urea denaturation test correctly identified the source of the elevated serum alkaline phosphatase (ALP) in 16 (64 percent). The serum gamma-glutamyl transpeptidase (GGTP) alone predicted the correct diagnosis in 64 percent of the cases and the urinary hydroxyproline (HOP) alone in 69 percent. When all three tests were performed simultaneously, the combination of results identified the source of the ALP in 88 percent of the cases. It is believed that this combination offers better sensitivity and specificity than any of the three tests used individually.


Formation of glycosidases in batch and continuous culture of Bacteroides fragilis. Nine strains of bacteroides fragilis were cultivated in stirred fermentors and tested for their ability to produce glycosidases. B. fragilis subsp. vulgatus B70 was used for optimizing the production of glycosidases. The highest bacterial yield was obtained in proteose peptone-yeast extract medium. The optimum pH for maximal bacterial yield was 7.0, and the optimum temperature for growth was 37 degrees C. The formation of glycosidases was optimal between pH 6.5 and 7.5, and the optimum temperature for synthesis of glycosidases was between 33 and 37 degrees C. Culture under controlled conditions in fermentors gave more reproducible production of glycosidases than static cultures in bottles. The strain was also grown in continuous culture at a dilution rate of 0.1 liter/h at pH 7.0 and 37 degrees C with a yield of 2.0 mg of dry weight per ml in the complex medium. The formation of glycosidases remained constant during the entire continuous process.


Clear, defined medium for the sporulation of Clostridium perfringens. A new, defined medium for the sporulation of Clostridium perfringens is presented. Sporulation levels exceeding 10(6) to 10(7) heat-resistant spores per ml were obtained for seven strains: PS49, PS52, FD-1, T-65, NCTC strains 8798, 8238, and 10240. In the presence of theophylline, a methylxanthine, higher levels of heat-resistant spores were attained for strains PS49, PS52, FD-1, ant T-65; photomicrographs demonstrated a higher fraction of sporulating cells when these strains were grown in the presence of methylxanthines. Use of washed, highly diluted (less than 100 cells) inocula resulted in no reduction in spore yield. Strain KA3 grew well but sporulated poorly on this medium. The medium was clear and free of precipitate when small amounts (100 microgram/ml) of methylxanthine were incorporated.


Simple bacterial preservation medium and its application to proficiency testing in water bacteriology. A medium composed of nutrient broth, 1.8% boric acid, and 1% sodium chloride at pH 7.0 was shown to maintain the stability of Escherichia coli cultures for up to 10 days at room temperature. By using this preservation medium for preparing a simulated sample a successful proficiency test survey in water bacteriology was conducted.


Reduction of azo dyes by intestinal anaerobes. Reduction of seven azo dyes (amaranth, Ponceau SX, Allura Red, Sunset Yellow, tartrazine, Orange II, and methyl orange) was carried out by cell suspensions of predominant intestinal anaerobes. It was optimal at pH 7.4 in 0.4 M phosphate buffer and inhibited by glucose. Flavin mononucleotide caused a marked enhancement of azo reduction by Bacteroides thetaiotaomicron. Other electron carriers, e.g., methyl viologen, benzyl viologen, phenosafranin, neutral red, crystal violet, flavin adenine dinucleotide, menadione, and Janus Green B can replace flavin mononucleotide. These data suggest that an extracellular shuttle is required for azo reduction.


Concentration of poliovirus from tap water onto membrane filters with aluminum chloride at ambient pH levels. A method is described for the concentration of an enterovirus from large volumes of tap water by addition of small amounts of aluminum chloride to enhance virus removal by membrane filters. Tap water treated with 2 X 10(-5) M aluminum chloride showed a slight decrease in pH (less than 0.5), a slight increase in turbidity, and enhanced removal of poliovirus by membrane filters. Virus was quantitatively recovered by treating the filters with a basic buffer, and this eluate was reconcentrated to a small volume by adsorption to aluminum hydroxide flocs. Using these procedures, virus from 1,000 liters of water was reduced to a final eluate of 20 to 80 ml with a mean recovery of 70%.


Hypotensive and bradycardic effects of classical alpha-sympathomimetic drugs upon intravenous administration to pentobarbitone-anaesthetized rats. The effects of the classical alpha-sympathomimetic drugs naphazoline, tramazoline, xylometazoline, tetryzoline and oxymetazoline on blood pressure and heart rate of pentobarbitone-anaesthetized (75 mg/kg; i.p.) normotensive rats were studied following intravenous injections. With the aid of this experimental animal model a dose dependent decrease in mean arterial pressure and heart rate could be established for these agents, oxymetazoline excepted, for which only the bradycardia could be quantified. It is suggested that central alpha-adrenoceptors are involved in the mechanism of this cardiovascular inhibition in view of the most specific alpha-adrenoceptor-stimulating properties of these ddrugs. Additionally, the results indicate that the use of pentobarbitone as the anaesthetic enhances the hypotensive action of these agents, so that their pronounced peripheral vasopressive response can be overcome.


[Arterial malformations and Takayashu disease]. The authors report a recent series of 5 cases with Takayashu's disease and discuss the common pathological presentation of malformations of the arterial trunk, caused by disorderd development of the arterial system or by later segmental arrested development of the large trunks. The disease, which presents in the young adult as a diffuse inflammatory process, is probably auto-immune, but has its foundations in embryonic and foetal development.


[Action of a sodium lactate perfusion on the hemodynamic effects of beta-blockaders]. In 15 dogs who had been given propranolol and 5 humans who had been given Pindolol, an infusion of sodium lactate produced: 1) an increased in blood lactate levels and pH; 2) an increase in the cardiac index; 3) a state in which an infusion of isoproterenol (inactive on its own) could act as it would have done had the betaadrenergic receptors not been blocked. A perfusion of lactic acid which was sufficient to cause the variations in pH and blood lactate levels that are seen with exercise had no action on the heart rate nor on the cardiac index of 10 dogs treated with propranolol and could not re-establish the betastimulating properties of isoproterenol. Finally, alkalosis produced by an infusion of THAM in 13 dogs or sodium bicarbonate in 2 humans, changed neither the cardiac index nor the heart rate, nor the response to isoproterenol after a betaadrenergic blockade. Infusion of sodium lactate associated with isoproterenol could be used to combat the depressent effects of betablockers in patients with cardiac disorders.


Bacterial 2,3-butanediol dehydrogenases. Enterobacter aerogenes, Aeromonas hydrophila, Serratia marcescens and Staphylococcus aureus possessing L(+)-butanediol dehydrogenase produced mainly meso-butanediol and small amounts of optically active butanediol; Acetobacter suboxydans, Bacillus polymyxa and Erwinia carotovora containing D(-)-butanediol dehydrogenase produced more optically active butanediol than meso-butanediol. Resting and growing cells of these organisms oxidezed only one enantiomer of racemic butanediol. The D(-)-butanediol dehydrogenase from Bacillus polymyxa was partially purified (30-fold) with a specific activity of 24.5. Except NAD and NADH no other cofactors were required. Optimum pH-values for oxidation and reduction were pH 9 and pH 7, respectively. The optimum temperature was about 60 degrees C. The molecular weight was 100000 to 107000. The Km-values were 3.3 mM for D(-)butanediol, 6.25 mM for meso-butanediol, 0.53 mM for acetoin, 0.2 mM for NAD, 0.1 mM for NADH, 87 mM for diacetyl, 38 mM for 1,2-propanediol; 2,3-pentanedion was not a substrate for this enzyme. The L(+)butanediol dehydrogenase from Serratia marcescens was purified 57-fold (specific activity 22.3). Besides NAD or NADH no cofactors were required. The optimum value for oxidation was about pH9 and for reduction pH 4.5. The optimum temperature was 32-36 degrees C. The molecular weight was 100000 to 107000. The Km-values were 5 mM for meso-butanediol, 10 mM for racemic butanediol, 6.45 for acetoin, 1 mM for NAD, 0.25 mM for NADH, 2.08 mM for diacetyl, 16.7 mM for 2,3-pentanedion and 11.8 mM for 1,2-propanediol.


Significance of microbial contamination of stored cadaver kidneys. The importance of microbial contamination of cadaver kidneys was assessed in 83 consecutively stored and transplanted kidneys. Fourteen kidneys had a single positive culture during storage and five had multiple positive cultures. Only one postoperative infection could be traced to kidney contamination during storage (Candida wound infection). In three of 64 patients who received noncontaminated kidneys, posttransplant wound infections developed. No wound infections occurred in 35 patients who received prophylactic antibiotics, whereas four wound infections occurred in 48 patients without antibiotic coverage. It is concluded that, although microbial contamination of stored cadaver kidneys occurs commonly, it is not an important source of infection in renal transplant recipients.


The Grimonster symposium on the occasion of the 50th anniversary of the first lumbar sympathectomy. Under the auspices of the Belgian Surgical Society, a small group of American and European surgeons discussed in detail the place of lumbar sympathectomy in present day surgical practice. A consensus was reached concerning the physiologic effect, anatomical variations and indications for the operation. Conclusions included the questionable value of lumbar sympathectomy for treating intermittent claudication due to aortoiliac disease alone, its possible effectiveness when obliterative arterial disease is limited to the femoral popliteal segment, and its occasional beneficial effect when performed in conjunction with some reconstructive arterial operations.


Formation of 3,N4-ethenocytidine moieties in RNA by vinyl chloride metabolities in vitro and in vivo. Rats were exposed to [1,2-14C] vinyl chloride. Liver RNA was isolated, hydrolyzed, and the nucleosides separated on Aminex-A-6. Besides the physiological bases and 1,N6-ethenoadenosine, radioactivity was also incorporated into 3,N4-ethenocytidine. Radioactive 3,N4-ethenocytidine moieties were also formed on incubation of polycytidylic acid with rat liver microsomes, NADPH and [14C] vinyl chloride. These alkylation mechanisms are consistent with the mutagenic and cancerogenic properties of vinyl chloride.


Studies of rat alkaline phosphatase I. Development of methods for detecting isoenzymes. A manual system of various estimations of rat plasma alkaline phosphatase activity has been devised for small volumes of plasma which uses different inhibitors, compares the utilisation of two substrates and includes acrylamide gel electrophoresis. The different inhibitors etc. allow a degree of discrimination between alkaline phosphatase extracts of rat organs. The properties of isoenzymes, e.g. intestinal phosphatase, differ depending upon the environment in which they are studied. In conjunction, if necessary, with the methods described for the estimation of liver and intestinal alkaline phosphatase activity, it is hoped to use the system to discriminate between the isoenzymes present in the plasma alkaline phosphatase of rats in toxicological studies.


[Experimental studies of the specificity and aspecificity of a beta-adrenolytic agent, bunitrolol (Kö 1366)]. In basis of our experimental researches bunitrolol presented: a) beta-adrenolytic activity prevailingly at cardiac level (superior to propranolol and practolol) in respect to vascular, tracheal, bronchial, uterus, intestinal and metabolic level; b) beta-adrenergic intrinsic activity; c) local anesthetic activity; d) chinidino-like activity; e) antiarrhythmic activity; f) hypotensive activity with interferences at the level of vasomotor centres, of the barosensitive zones, of the renin secretion and of the purinergic periphery.


[Experimental analysis of the cardiovascular effects of acebutolol, a drug with beta-adrenolytic activity]. In experiments at cardiovascular system level, acebutolol was found to possess beta-adrenolytic activity of cardioselective type associated with intrinsic beta-adrenergic, antiarrhythmic and anti-hypertensive activity. It was well tolerated systemically.


Local injuries by accidental ingestion of corrosive substances by children. Own data and analysis of previous publications show that situations where accidental ingestion of corrosive substances by children may have happened are frequent, but severe corrosive esophagitis leading to perforation or stricture formation is very rare. In case of suspected esophageal injury, esophagoscopy and glucocorticoid treatment become necessary. The evaluation of the initial symptoms in patients from our own material and from the literature indicates that all children with serious esophageal burns had one or more of the following symptoms: visible burns in the oral cavity, hypersalivation, retching, vomiting, retrosternal or epigastric pain, cardiovascular collaps, airway stenosis. Hence, children with an uncertain history of ingestion and without any of these symptoms need not be treated. After ingestion of liquid substances, but never of dry or granular products, lesions in the esophagus without accompanying burns in the oral cavity were observed. The evaluation of 1158 cases of accidental ingestions of several types of household products and a collection of data from the literature on the causticity of these substances shows that cleaners containing mainly detergents and phosphates (with pH values generally between 9 and 11), and household bleaches on sodium hypochlorite basis, are relatively harmless. Drain cleaners (NaOH), decalcifiers (formic acid) and detergents for automatic dish washing machines (metasilicates) are very caustic and are responsible for the majority of serious accidents in children.


Poly(adenosine dephosphate ribose) metabolism and regulation of myocardial cell growth by oxygen. Control of the rate of cardiac cell division by oxygen occurs most probably by altering the redox state of a control substance, e.g. NAD(+)right harpoon over left harpoonNADH. NAD(+) (and not NADH) forms poly(ADP-ribose), an inhibitor of DNA synthesis, in a reaction catalysed by poly(ADP-ribose) polymerase. Lower partial pressure of oxygen, which increases the rate of division, would shift NAD(+)-->NADH, decrease poly(ADP-ribose) synthesis, and increase DNA synthesis. Chick-embryo heart cells grown in culture in 20% O(2) (in which they divide more slowly than in 5% O(2)) did exhibit greater poly(ADP-ribose) polymerase activity (+83%, P<0.001) than when grown in 5% O(2). Reaction product was identified as poly(ADP-ribose) by its insensitivity to deoxyribonuclease, ribonuclease, NAD glycohydrolase, Pronase, trypsin and micrococcal nuclease, and by its complete digestion with snake-venom phosphodiesterase to phosphoribosyl-AMP and AMP. Isolation of these digestion products by Dowex 1 (formate form) column chromatography and paper chromatography allowed calculation of average poly(ADP-ribose) chain length, which was 15-26% greater in 20% than in 5% O(2). Thus in 20% O(2) the increase in poly(ADP-ribose) formation results from chain elongation. Formation of new chains also occurs, probably to an even greater degree than chain elongation. Additionally, poly(ADP-ribose) polymerase has very different K(m) and V(max.) values and pH optima in 20% and 5% O(2). These data suggest that poly(ADP-ribose) metabolism participates in the regulation of heart-cell division by O(2), probably by several different mechanisms.


Intestinal neuraminidase activity of suckling rats and other mammals. Relationship to the sialic acid content of milk. 1. The neuraminidase activity of homogenates of the mucosa of the middle and distal thirds of the small intestine of rats increased about 5-fold between birth and 4 to 8 days of age, and then gradually declined to the much lower adult activity by 24 days. No comparable changes occurred in the proximal third. 2. In 8-day-old rats, the neuraminidase activity of the middle and distal thirds of the small intestine was about 10 times greater than that of the proximal third, 20 times greater than that of the colon and at least 100 times greater than that of the liver, brain, gastric mucosa or pancreas. 3. In all other species investigated (mice, rabbits, cats and guinea pigs), the neuraminidase activity of the middle and distal thirds of the small intestine was greater in suckling animals than in adults. 4. The sialic acid content of rat milk increased about 2-fold between birth and 8 days post partum and then declined. 5. There was a highly significant positive correlation between the intestinal neuraminidase activity of suckling animals of various species and ages and the sialic acid content of milk obtained from the corresponding species and stage of lactation. 6. It is suggested that the intestinal neuraminidase of suckling mammals functions primarily to remove sialic acid from various components of milk, thus providing sialic acid for the synthesis of sialoglycoproteins and gangliosides by the young.


[Reaction kinetic studies on the formation of n-nitrosoephedrine in vitro and in vivo (author's transl)]. N-Nitrosoephedrine was synthesized at a high yield (greater than 90%). The influence of pH on the reaction process (ephedrine sulfate + nitrous acid in hydrochloric acid) was investigated and the pH-optimum determined. Furthermore the influence of coffee, ascorbic acid, citric acid, cyclamate/saccharin (10:1), smoke condensate of cigarettes, glucose and ethanol on the reaction process was also determined. The tests with animals (rats) showed that N-nitrosoephedrine could be detected in serum after the oral application of ephedrine sulfate and sodium nitrite.


Clinical experiments with the new oral long-acting neuroleptic clopimozide (R 29 764). A pilot study with the new oral long-acting neuroleptic agent clopimozide, a diphenylbutylpiperidine derivative, has been reported. 30 female patients, only 20 of whom completed the trial, were treated with dosages of between 5 and 35 mg. The treatment comprised four periods in which the maximum tolerated daily dosage, the optimal weekly dosage (mean: 20.25 mg), the maximum tolerated weekly dosage and the optimal daily dosage were to be determined. In 10 patients plasma levels were determined with the radio-immunological method. The product exhibited a marked effect on schizophrenic target symptoms (significant at the 0.001 and 0.01 levels), of at least one week's duration, with an absence of sedation and a low degree of extrapyramidal side-effects. Clopimozide appears to be particularly suitable for the neuroleptic long-term treatment of compensated schizophrenic patients with insight into their disease.


Beta-blockers and asthma.

Beta-blockers and asthma. In a single-blind, randomised, crossover study in 10 asthmatic patients, the effects of approximately equipotent oral doses of 3 cardioselective beta-blockers-atenolol (100 mg), metoprolol (100 mg), and acebutolol (300 mg)-and 4 non-cardioselective beta-blockers-proranolol (100 mg), oxprenolol (100 mg), pindolol (5 mg), and timolol (10 mg) upon FEV1 were compared. All drugs, except pindolol, produced a significant reduction in standing pulse rate and prevented an increase in heart rate after inhaled isoprenaline (1500 microgram). All drugs caused a fall in FEV1 but only atenolol did not differ significantly from placebo in this respect. The bronchodilator response to inhaled isoprenaline was blocked by the 4 non-cardioselective drugs; the 3 cardioselective agents permitted some bronchodilatation, but only atenolol did not differ from placebo.


Comparison of diazepam and flunitrazepam for sedation during local anaesthesia for bronchoscopy. Diazepam and flunitrazepam were compared as amnesic and sedative adjuncts to local anaesthesia for diagnostic bronchoscopy in 92 patients. After local anaesthesia of the pharynx, larynx and trachea with lignocaine, atropine plus diazepam of flunitrazepam was injected i.v. The co-operation of the patients and the technical circumstances under which the bronchoscopy was performed were good in each group. None of the treatments significantly modified arterial pressure or heart rate. Two hours after the injection, flunitrazepam 0.01 mg kg-1 more frequently caused amnesia for pictures shown to the patients during the first 15 min after injection (failure to recall 42--75%, and for bronchoscopy 67%), than did diazepam 0.125 mg kg-1 (failure to recall 21--67%; bronchoscopy 38%). Double doses of the drugs caused amnesic actions similar to those of flunitrazepam 0.01 mg kg-1. When failure to recall was assessed on the following day, 29% and 5% of the patients remembered bronchoscopy after flunitrazepam 0.01 and 0.02 mg kg-1 respectively; after diazepam 0.125 and 0.25 mg kg-1 the corresponding percentages was 59% and 30% (P less than 0.05% v. fluintrazepam). The ability to stand and walk on a stright line was similar after the smaller doses of both drugs, but after the larger doses recovery was slower after flunitrazepam. Psychomotor performance was still distinctly impaired 2 h after the injection of the larger doses.


Fazadinium in anaesthesia.

Fazadinium in anaesthesia. The clinical use of the neuromuscular blocking agent fazadinium was investigated in a survey of 500 patients. Rapid conditions for tracheal intubation followed by excellent relaxation and easy antagonism were obtained. It is suggested that these findings commend it as an alternative to suxamethonium. Tachycardia appeared to be the only disadvantage associated with its use.


Acid hydrolases in blister fluid. I. Characterization and quantification of acid phosphatase. Acid phosphatase has been characterized and quantified in human suction blister fluid, interstitial fluid and serum. The acid phosphatases of suction blister fluid and serum showed differences in their pH activity curves, Michaelis-Menton constants, heat stabilities and sensitivities to inhibition by tartrate and fluoride. The behaviour of the interstitial fluid enzyme was intermediate between these two. The levels of activity in blister fluid and serum were very similar, both being more than twice that of interstitial fluid. These results suggest that the acid phosphatase activity in suction blister fluid is derived largely from the overlying epidermis rather than from serum.


Acid hydrolases in blister fluid. II. Characterization and quantification of glycoside hydrolases. Seven glycoside hydrolases have been investigated in suction blister fluid, interstitial fluid and in serum. Six of these have been characterized; no differences could be demonstrated between the corresponding enzymes from the various sources. The remaining enzyme (beta-glucosidase) was not found. Quantitative data suggest that 2 enzymes (beta-acetylglucosaminidase and beta-glucuronidase) diffuse freely from the epidermis into blister fluid, whereas 4 (alpha-glucosidase, alpha- and beta-galactosidase and alpha-mannosidase) are almost entirely retained in the roof of the bulla.


Metabolic changes associated with intake by cows of complete diets containing straw and concentrates in different proportions. 1. Pelleted diets containing concentrates and 0, 200, 400 or 600 g chopped straw/kg were fed ad lib, for 5 h daily to four cows. Concentrations of various energy-yielding metabolites were measured in samples of rumen fluid and jugular blood taken before feeding and at intervals after food was offered. 2. After feeding, the pH of rumen fluid decreased rapidly and the total concentration of volatile fatty acids (VFA) increased; the changes were greatest in the diet containing no roughage. The changes were essentially complete by 135 min at approximately the time feeding stopped. Rumen lactate concentrations were always low and increased significantly only in cows given no roughage. 3. There were highly significant relationships between the peak rumen acetate concentration after a meal and the apparent digestibility of the dry matter of that meal, and the amount of material in the rumen at the end of a meal. The latter relationship resulted in there being no difference between treatments in the total amount of acetate present in the rumen after feeding. 4. In the blood, concentrations of acetate, propionate, beta-hydroxybutyrate (BHB) and lactate all increased after feeding. The increase was prolonged and maximum values were rarely reached before 4-5 h. The highest concentrations of acetate and BHB were found in cows given 20 g roughage/kg and were twice as great as those in cows given no roughage; lower concentrations were found in cows given 400 or 600 g roughage/kg. This information, together with the rumen concentrations of acetate and butyrate, was interpreted as indicating an inhibition of VFA absorption from the rumen of cows given no roughage. 5. Plasma glucose concentrations decreased rapidly for 4 h after feeding, the decrease being greatest in cows given 200 g roughage/kg. Non-esterified fatty acid concentrations also decreased after feeding 200 and 0 g roughage/kg rations, but concentrations were not high at any time. 6. It is concluded that chemical changes in the rumen correspond to feeding behaviour much more closely than changes in blood and therefore any chemostatic regulation of food intake probably occurs at the former site, with acetate playing a major role. Monitoring energy balance at tissue level is likely to be mediated hormonally, with insulin an important factor.


On the measurement of pH in Escherichia coli by 31P nuclear magnetic resonance. The 31P high resolution NMR spectra of concentrated suspensions of Escherichia coli cells have been measured at 145.8 MHz. The position of the orthophosphate resonance is used as a measure of internal and external pH. In accord with Paddan, Zilberstein and Rottenberg ((1976) Eur. J. Biochem. 63, 533--541) it is shown that when properly energized the internal pH is 7.5 +/- 0.1. By synchronizing the NMR data acquisition with 3-s bursts of O2 it is possible to measure the internal pH with a time resolution of about 1 s. It is shown that at 20 degrees C the pH remains constant for times longer than 15 s after the oxygen is discontinued and it decays in several minutes.


A study of H+ transport in gastric microsomal vesicles using fluorescent probes. Fluorescent amines, 9-aminoacridine, acridine orange and quinacrine, were used as probes for a pH gradient (deltapH) across gastric microsomal vesicles. Analysis of probe uptake data indicates that 9-aminoacridine distributes across the membrane as a weak base in accordance with the deltapH. On the other hand, acridine orange and quinacrine show characteristics of binding to membrane sites in addition to the accumulation in response to deltapH. A discussion of the advantages and limitations of the probes is presented. Application of these probes to pig gastric microsomal vesicles indicates that that K+-stimulated ATPase is responsible for the transport of H+ into the vesicles and thus develops a deltapH across the membrane. The deltapH generated by the K+-ATPase has a definite requirement for internal K+. The proton gradient can be discharged slowly after ATP depletion or rapidly either by detergent disruption of the vesicles or by increasing their leakiness using both H+ and K+ ionophores. On the other hand, the sole use of the K+ ionophore, valinomycin, stimulates the ATP-induced formation of deltapH by increasing the availability of K+ to internal sites. This stimulation by valinomycin requires the presence of permeable anions like Cl-. Analysis of the Cl- requirement indicates that in the presence of valinomycin the net effect is the accumulation of HCl inside the gastric vesicles. With an external pH of 7.0, the ATP-generated deltapH was calculated to be from 4 to 4.5 pH units. The results are consistent with the hypothesis that the K+-stimulated ATPase drives a K+/H+ exchange across the gastric vesicles. Since other lines of evidence suggest that these gastric microsomes are derived from the tubulovesicular system of the oxyntic cell, the participation of the ATP-driven transport processes in gastric HCl secretion is of interest.


NMR kinetic studies of the ionophore X-537A-mediated transport of manganous ions across phospholipid bilayers. Nuclear magnetic resonance spectroscopy has been applied as a method for studying manganous ions transport across the membrane of phosphatidylcholine vesicles. The rates of the ionophore X-537A (lasalocid A)-mediated Mn2+ transport have been measured as a function of ionophore concentration, pH of the vesicle suspension, and temperature. The translocation was found to occur via a neutral complex composed of one manganous ion bound in two ionized X-537A molecules (Mn X2). The activation energy for the overall transport process was determined to be 22 +/- 5 kcal/mol. Also a pKa of 5.0 +/- 0.2 was determined for the ionophore acid dissociation equilibrium in the vesicle suspension.


Effects of pH on beta-hydroxybutyrate transport in rat erythrocytes and thymocytes. Entry of beta-hydroxybutyrate into erythrocytes and thymocytes is facilitated by a carrier (C), as judged from temperature dependence, saturation kinetics, stereospecificity, competition with lactate and pyruvate, and inhibition by moderate concentrations of methylisobutylxanthine, phloretin, or alpha-cyanocinnamate. We studied the dependence of influx and efflux on internal and external pH and [beta-hydroxybutyrate]. Lowering external pH from 8.0 to 7.3 to 6.6 enhanced influx into erythrocytes by lowering entry Km from 29 to 16 to 10 mM, entry V being independent of external pH. Lowering external pH inhibited efflux. At low external pH, external beta-hydroxybutyrate enhanced efflux slightly. At high external pH, external beta-hydroxybutyrate inhibited efflux. Internal acidification inhibited influx and internal alkalization enhanced influx. Internal beta-hydroxybutyrate (betaHB) enhanced influx more in acidified than alkalized cells. These data are compatible with coupled betaHB-/OH- exchange, betaHB- and OH- competing for influx, C:OH- moving faster than C: betaHB-, empty C being immobile. They are also compatible with coupled betaHB-/H+ copermeation, empty C moving inward faster than H+:C:betaHB-, H+:C being immobile, and C:betaHB- (without H+) being so unstable as not to be formed in significant amounts (relative to C, H+:C, and H+:C:betaHB-).


The kinetics and equilibria of squirrel-fish hemoglobin. A Root effect hemoglobin complicated by large subunit heterogeneity. The functional properties of squirrel-fish hemoglobin have been measured by studying ligand binding equilibria and kinetics. The results show that squirrel-fish hemoglobin has a Root effect with a corresponding stabilization of the low affinity state. The properties of this state are pH dependent even in the absence of cooperativity. The effect of ATP shifts the overall ligant affinity towards the low affinity state and is characteristic of the allosteric effect caused by organic phosphates. Under pH and ATP conditions favoring the low affinity conformational state, a 10-fold difference in the binding kinetics of carbon monoxide to the alpha and beta subunits is observed.


Circular dichroism of metallothioneins. A structural approach. A comprehensive study on circular dichroism of metallothioneins containing Zn, Cd and Cu was carried out. The contributions of the metals, the sulphur and the polypeptide chain to the observed Cotton effects was shown. From the pH dependency of the extrinsic Cotton effects which are due to the metal-thiolate chromophore the stability of the metal clusters was found to decrease in the order Cu greater than Cd greater than Zn. The pH values corresponding to the dissociation of half of the bound metal ions are 0.44 for Cu-thionein, 3.05 for Cd-thionein and 4.6 for Zn-thionein. The extrinsic Cotton effects of Cd, Zn-thioneins of varying Cd to Zn ratio could be simulated using the difference circular dichroic spectra of Cd-thionein (bands at 227, 242.5 and 262 nm), Zn-thionein (bands at 225 and 244 nm) and the circular dichroic spectrum of cysteine-thionein (band at 200 nm, shoulder at 225 nm). Since during the dissociation of the metals the circular dichroic spectra exhibited changes only in amplitude and not in shape we can conclude that the dissociation of the metal ions involves the complete sequential degradation of metal clusters. In the near-ultraviolet region the metal-free proteins show only Cotton effects attributable to a disulphide chromophore. Thus Cotton bands are observed for cystine-thionein at 282.5 and 260 nm. From the intrinsic circular dichroism of Cd- and Zn-thionein (negative Cotton effect at 200 nm, shoulder at 225 nm) it follows that the protein conformation consists of less than 5% helical or pleated sheet structure and therefore has to be classified as unordered structure or "fixed" random coil


Changes in structure and hydrophobic surface properties of beta-lactoglobulin determined by partition in aqueous two-phase polymeric systems. The non-polar surface properties of beta-lactoglobulin and especially its interaction with poly(ethylene glycol)-bound palmitate has been studied as a function of pH, temperature and protein concentration. The maximum interaction between beta-lactoglobulin and polymer-bound palmitate occurs at pH 4.3 and pH 7.8. The change in conformation of beta-lactoglobulin around pH 7.5 seems to involve exposure of apolar amino acids to the solvent which results in an increased affinity for hydrocarbons. This is contrary to the situation at pH 4.8--6.0 where the corresponding change in conformation does not affect the protein-hydrocarbon interaction. The results suggest that partition studies in an aqueous two-phase system is a very useful tool to detect changes in conformation and aggregation and to characterize the corresponding hydrophobic surface properties of a protein.


Influence of phosphate ligands in abolishing the conformational difference between ribonuclease A and its acid-denatured derivative. The initial structural alteration of RNAase A due to acid denaturation (0.5 N HCl, 30 degrees C) that accompanies deamidation (without altering enzymic activity) has been dectected by spectrophotometric titration, fluorescence and ORD/CD measurements. It is shown that acid treated RNAase A has an altered conformation at neutral pH, 25 degrees C. This is characterized by the increased accessibility of buried tyrosine residue(s) towards the solvent. The most altered conformation of RNAase A is found in the 10 h acid-treated derivative. This has about 1.5 additional exposed tyrosine residues and a lesser amount of secondary structure than RNAase A. All three methods (titration, fluorescence and CD) established that the structural transition of RNAase A is biphasic. The first phase occurs within 1 h and the resulting subtle conformational change is constant up to 7 h. Following this, after the release of 0.55 mol of ammonia, the major conformational change begins. The altered conformation of the acid-denatured RNAase A could be reversed completely to the native state through a conformational change induced by substrate analogs like 2'- or 3'-CMP. Thus the monodeamidated derivative isolated from the acid-denatured RNAase A by phosphate is very similar to RNAase A in over-all conformation. The results suggest the possibility of flexibility in the RNAase A molecule that does not affect its catalytic activity, as probed through the tyrosine residues.


Physicochemical properties of cartilage proteoglycans extracted by lanthanum chloride. Bovine nasal cartilage was extracted with 0.5 M LaCl3 and the extract then diluted with nine volumes of water. The resulting precipitated (PLaCl3) contained the proteoglycan subunits, together with minor protein components, but was essentially free from hyaluronic acid. The properties of PLaCl3 were investigated by chemical analysis, electrophoresis, viscometry and analytical ultracentrifugation, and the results compared with those for proteoglycan obtained by caesium chloride density gradient centrifugation of 2 M CaCl2 cartilage extracts. Proteoglycan subunits (A1D1) prepared from PLaCl3 showed identical properties to those obtained from other high ionic strength cartilage extracts.