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In congenital blindness, the absence of vision can be compensated for by the utilization of the tactile and auditory senses, resulting in normal affect and ego development. Some of the major problems in the way of such compensation were noted. Adventitious blindness produces disruption of ego functions, with severe affect manifestations, notably grief and mourning reactions and actual dependency. The factors favoring successful or unsuccessful resolution of mourning and restoration of normal ego functions were indicated.

Comparisons of the nests of associations within which a single memory reappeared during the course of an analysis deemed successful by analysis and patient revealed changes in content and structure of these matrices. Judges, given a sample of associations surrounding a reappearance, were able to relate a particular appearance to its sequential location in the development and flow of an analysis. In the study samples, the patient became increasingly active and insightful as the analysis progressed. Resistance decreased, and the memory, initially given as a narrative report, became a powerful metaphor for the examination of conflicts concerning aggression and its defense, passivity. The results of the judges' analyses and of the thematic analysis suggest that a close examination of associations surrounding a repeated stimulus, a single memory, can be used as an index of analytic progress.

We have described some of the problems encountered in an analyzability committee's evolving approach to the assessment of applicants to a psychoanalytic clinic. A primary error was to think in terms of the level of drive development--equating pathology at an oedipal level with analyzability. Recognition that the more important issue for assessment purposes was the presence or absence of those ego capacities and personality characteristics necessary for success in analysis has led to a reorientation of our approach.

The relative iron, copper and mercury contents of individual, isolated erythrocytes from eight people were determined by analytical electron microscopy. The variation in iron content between erythrocytes of the same sample is more than six times, for copper content more than tem times and for mercury more than five times. Similar variations were observed for 1-day-old chick defintive erythrocytes and for 4-day-old chick embryo primitive erythrocytes. The range of variation does not depend greatly, it at all, on the age of the erythroyctes or the tissue of origin. There is little or no correlation between the variation of iron contnet and that of copper. The cause of the wide variation of metallic ion contnt among erythroyctes is not yet known.

A technique of hypophysectomy and regimes of pre- and post-operative care were developed for the tammar wallaby, Macropus eugenii, to a stage when animals can survive the operation with little apparent stress. Thyroid and adrenal gland weights declined after hypophysectomy, especially within the first 20-30 days. Changes in the adrenal cortex after hypophysectomy suggested that this region may have a zonal organization different from that in eutherian mammals. The reproductive tracts of males and females lost weight rapidly after hypophysectomy. Eleven plasma parameters were studied for the effects of hypophysectomy. There was a reduction in sodium and chloride and a tendency to higher potassium levels, reflecting inadequate adrenal cortical function. Calcium and total protein values remained unaffected, but inorganic phosphate, glucose and cholesterol were depleted, and a less significant depletion in blood urea, nitrogen and uric acid was evident.

Intact, breeding female budgerigars enter and occupy a nestbox 8-10 days before egg-laying. Ovariectomized budgerigars did not enter nestboxes. Oestradiol-17 beta monobenzoate (OB) (0-05 mg, i.m.) induced ovariectomized birds to enter nestboxes. Higher dosages (0-1 or 0-5 mg) did not facilitate this effect. The amount of time spent in nesting behaviour by all OB-treated groups did not differ from that shown by a group of intact females during the initial phase of the nesting sequence (first 4 days in the nestbox), but was significantly less than that of intact females undergoing later stages of nesting behaviour (4 days before and after laying the first egg). Oestradiol induced a increase in oviduct weight and size which was dose-dependent. With a low dose (0-05 mg OB), the oviducts were slightly larger than those of intact females which had been in the nestbox 1-2 days. With higher dose levels (0-1-0-5 mg OB), precursor albumen granules had formed in tubular glands of the magnum, a stage more typical of females which had been in the nestbox 4-6 days. The results indicate that low levels of oestrogen induce female budgerigars to enter nestboxes and initiate oviduct development. Subsequent phases of the nesting behaviour and oviduct development may be causally related to other hormones.

Photorefractory intact female canaries were exposed to a photoperiod of 8 h light: 16 h darkness (8L: 16D) for 0, 2, 4 or 6 weeks before photostimulation with 14L: 10D for 4 weeks. Photoresponsiveness was measured in terms of plasma immunoreactive LH, ovary and oviduct weights, follicle size and nest-building behaviour. Birds not exposed to 8L: 16D were unresponsive to photostimulation while those exposed for 6 weeks to 8L: 16D were as fully photosensitive as females similarly treated immediately before their first breeding season. Birds exposed for 2 or 4 weeks to 8L: 16D responded to subsequent photostimulation but were not as responsive as birds exposed to short days for 6 weeks. Positive correlations between the two ovarian parameters and between these ovarian parameters and oviductal weight existed in all groups exposed to short days, but not in birds not exposed to short days. It is concluded that the restoration of photosensitivity at the end of the refractory period is a function of the number of short days experienced; about 6 weeks of short days are needed for its completion. In another experiment photorefractory birds exposed for 6 weeks to short days without subsequent photostimulation, although potentially photosensitive, were indistinguishable (in terms of the above parameters) from refractory birds kept on long days for the same period; in these birds refractoriness was not broken.

The effect of alpha-MSH on sebum secretion and preputial gland weight was examined in intact, castrated and hypophysectomized male rats and in hypophysectomized rats receiving treatment with either testosterone propionate (TP) or progesterone. After treatment with alpha-MSH for 2 weeks, increases in sebum secretion occurred in intact, castrated and hypophysectomized rats, but larger responses were found in the hypophysectomized rats that had received treatment with either TP or progesterone, suggesting that alpha-MSH acts synergistically with TP and progesterone to stimulate sebum secretion. Alpha-Melanocyte-stimulating hormone also increased preputial gland weight in intact rats, but there was no response after castration and only a small response after hypophysectomy. However, when the hypophysectomized rats received simultaneous treatment with either TP or progesterone, alpha-MSH increased preputial gland weight. It is suggested that alpha-MSH acts directly on the sebaceous glands to stimulate lipogenesis and, together with steroid hormones, may have an important role in controlling sebaceous glandd function in the rat and other hairy mammals. With the evolution of hair, certain of the MSH peptides may have lost their significance as pigmentary hormones and have developed a sebotrophic function. For this reason, it might be more appropriate to refer to these peptides as the 'sebotrophins'.

The concentration of LH in the plasma of guinea-pigs from day 50 of gestation to day 45 of postnatal life was assayed by radioimmunoassay utilizing a cross-reaction with anti-ovine LH antiserum. The effect of gonadectomy in infancy and in the adult upon the plasma concentration of LH was also studied. The LH concentration in the plasma of male or female foetuses was high immediately prenatally and fell at birth. High levels of LH were again detected in male, with a lesser increase in female, guinea-pigs over the first 10 days postnatally. Maternal plasma concentrations of LH remained consistently low. Removal of the gonads on days 0, 5, 10, 15, 25 or 35 of postnatal life, followed by blood collection at autopsy 10 days later, caused a significant rise in plasma LH content at all ages. The rise in plasma LH after gonadectomy in adults was less marked in male than in female guinea-pigs.

Binding of 5alpha-androstane-3alpha, 17 beta-diol (3alpha-diol) and 5alpha-androstane-3beta,-17 beta-diol (3beta-diol) in vivo and in vitro to the 100 000 g cytosol fraction of the rat prostate and seminal vesicles as well as to plasma was studied by agargel electrophoresis and sucrose density gradient ultracentrifugation and the results compared with the corresponding findings for 5alpha-dihydrotestosterone (5alpha-DHT). The metabolism of 3alpha-diol and 3beta-diol was also investigated by thin-layer chromatography. The following results were obtained: (1) A specific binding of 3alpha-diol and 3beta-diol by the cytosols could not be demonstrated in vitro, while 5alpha-DHT was specifically bound. (2) In plasma, 3alpha-diol was extensively bound, 3beta-diol less extensively bound, while 5alpha-DHT remained unbound. (3) After intravenous injection of 3alpha-diol, specifically bound radioactivity, increasing within 30 min, was found in the prostate cytosol, while after 3beta-diol injection no binding occurred. (4) Parallel to the increased binding, the total radioactivity in the prostate accumulated within 30 min after 3alpha-diol injection, the uptake being 5-3 times higher than in skeletal muscle. However after 3beta-diol injection, total radioactivity decreased in the prostate within 30 min, the uptake being only 1-5 times higher than in skeletal muscle. (5) One minute after injection of 3alpha-diol, 53% of the extracted radioactivity in the prostate had been converted to 5alpha-DHT, this increased within 30 min to 81%. Thirty minutes after the injection of 3beta-diol, about 32% of the extracted radioactivity in the prostate had been converted to 5alpha-DHT. (6) From the in-vivo and in-vitro experiments it was concluded that 3alpha-diol exerts its biological effects mainly by its conversion into 5alpha-DHT.

The concentrations of FSH, LH, prolactin, oestradiol and progesterone were measured in peripheral plasma and follicular fluid of women throughout the menstrual cycle. With the exception of prolactin, concentrations of pituitary and steroid hormones in follicular fluid correlated with those in peripheral plasma. Follicle-stimulating hormone was present in a greater number of small follicles (smaller than 8mm) during or just after the peaks of FSH in peripheral plasma. During the mid-follicular phase the concentration of both FSH and oestradiol in fluid from large follicles (larger than or equal to 8 mm) was high. During the late follicular phase the large follicles (larger than or equal to 8 mm) contained high amounts of progesterone in addition to oestradiol, low physiological levels of prolactin, and concentrations of LH and FSH about 30 and 60% respectively of those found in plasma. By contrast no large 'active' follicles (larger than or equal to 8 mm) were found during the luteal phase although many contained both LH and FSH. Luteinizing hormone was present in a proportion of small follicles (smaller than 8 mm) during the late follicular and early luteal but not at other stages of the menstrual cycle. It is suggested that a precise sequence of hormonal changes occur within the microenvironment of the developing Graafian follicle; the order in which they occur may be of considerable importance for the growth of that follicle and secretory activity of the granulosa cells both before and after ovulation.

After two daily ovine prolactin injections in rats, significant increases in jejunal absorption of glucose, glycine and proline, as well as of fluid and NaCl, occurred. Although ouabain caused dramatic reductions in fluid, sodium, chloride and glucose transport, prolactin had an effect even in the presence of ouabain. Mucosal hypersomolaity significantly decreased fluid, sodium and chloride absorption but had no significant effect on glucose transport. Addition of prolactin in vitro had no effect on intestinal absorption. Prolactin-induced increases in transport of fluid, NaCl and organic nutrients by the mammalian jejunum may play an important role in nutritional and osmoregulatory adaptations.

The fine structure of the zona pellucida of mouse unfertilized egg cells and of mouse embryos was investigated. In addition to standard fixation for electron microscopy, fixation with an addition of ruthenium red was used and found to be especially valuable for the purpose of studying the zona pellucida. The zona pellucida of unfertilized egg cells and of embryos consisted mainly of fibrillar and granular material. There were two layers: a thicker internal and a denser external layer in the zona pellucida of the unfertilized egg cells. The zona pellucida of embryos had, in addition to these two layers, a coarse-grained intimal layer inside the internal layer and a fine-grained peripheral cortical reaction whereas the peripheral layer can be accumulated and modified during the passage of the embryo through the female genital tract. The influence of developmental events and environmental factors on the fine structure of zona pellucida was discussed.

The ability of caput, corpus, and cauda epididymal and vasa deferentia mouse sperm to fertilize eggs in vitro was investigated. Cauda epididymal sperm fertilized 45 of 54 ova, whereas caput and corpus sperm fertilized a negligible number of ova, thus indicating their immaturity. Vasa deferentia sperm fertilized only 10 of 52 ova and their reduced fertility may be due to aging in vivo. No difference in preimplantation development of the resulting zygotes with respect to the sperm source was observed. Three centrifugations and resuspensions of cauda epididymal sperm in fresh medium did not affect their fertilizing ability. I conclude that fertilization in vitro can occur after removal of epididymal secretions.

Elimination of 137-Cs and 59-Fe by three species of wild rodents was measured in laboratory and field experiments to determine whether excretion rates of these nuclides are influenced directly by general metabolic (CO-2 production) rates. Sigmodon hispidus and Peromyscus leucopus were used in both field and laboratory experiments, and Reithrodontomys humulis was investigated only in the laboratory. Final-component biological half-lives (Tb) of 59-Fe for Sigmodon averaged 108 days in winter, 144 days in spring, and means ranged from 176 to 242 days under various laboratory conditions (ambient temperature, cold-exposure, irradiation, and chemical metabolic inhibition); for Peromyscus, values were 50 days in winter 47 days in spring, and 289 days (at ambient temperature only) in the laboratory; for Reithrodontomys mean values ranged from 121 to 178 days at different laboratory treatment levels. Biological half-lives of final-component 137-Cs elimination for Sigmodon averaged 7.5 days in winter, 7.9 days in spring, and means ranged from 7.7 to 8.6 days at the different treatment levels in the laboratory. Laboratory mean values for Reithrodontomys ranged from 3.5 to 3.9 days. For Peromyscus, Tb values averaged 3.4 days in winter, 3.6 days in spring, and 3.5 days in the laboratory. The data suggested that elimination of 59-Fe is influenced by metabolic rates of rodents in the field, but laboratory experiments were unable to demonstrate any predictable relationship. Neither did the rate of final-component 137-Cs loss from rodents appear to be influenced by metabolic rate in the laboratory or in the field. However, final-component Y-axis intercept values of 137-Cs exhibited a linear correlation with metabolic rates, and equations were derived from these intercept values to predict metabolism in the field for two species: for Sigmodon, Daily Average CO-2 Production (ml/hr/g, STP) [5.24-0.9172 (log-ea)] [0.886]; and for Reithrodontomys, = [12.51-1.80 (log-ea)] [0.886].

The presence of microfibrillar structures in the nucleus and cytoplasm of Amoeba proteus has been described after glutaraldehyde and osmium fixation. The possible roles of cytoplasmic microfibrils in the contraction process of amoeba and nuclear microfibrils in the formation of the honeycomb nuclear lamina are discussed.

UV irradiation of the vegetal pole of stage 2 X. laevis embryos with doses of 6000-18000 ergs/mm-2 delayed or inhibited cytokinesis in the vegetal hemisphere. Nuclear division continued so that a syncytium was formed, the size and persistence of which depended on the dose. Embryos which received a low UV dose were subsequently able to recover whereas embryos receiving a high dose were unable to gastrulate successfully. The implications for investigations into the role of cytoplasmic determinants are discussed.

In X. laevis embryos injected with (3-H) thymidine between early gastrula and late neurula, the presumptive primordial germ cell nuclei were labelled and some were observed in mitosis, confirming that division occurs throughout this developmental period. In all labelled and mitotic cells, the germ plasm adjoined the nucleus, an observation which invalidates the hypothesis that germ plasm inhibits mitosis in presumptive primordial germ cells after gastrula. In embryos which had been irradiated with UV at the vegetal pole at the 2-cell stage, the presumptive primordial germ cells also incorporated (3-H) thymidine and divided during the same developmental period.

Physical and nutritional factors required for growth of Bacteroides nodosus isolates from ovine foot-rot lesions were examined. Simplified anaerobic culture techniques were devised utilizing a fully soluble, autoclavable, liquid medium (TAS) which contained proteose-peptone, yeast and meat extracts and certain other essential compounds required to promote prompt and serially transferrable growth of cultures from small inocula. The latter included Trypticase, arginine, a reducing agent (most suitably thioglycollic acid) and CO2; serine and Mg2+ markedly increased growth yields. Trypticase could not be replaced by a commercial preparation of acid-hydrolysed casein; other forms of hydrolysed protein gave delayed and inconsistent growth. Maximum growth of cultures required concentrations of 0-02 to 0-35 M-arginine, which could not be replaced by glutamic acid, citrulline or ornithine. Exogenous carbohydrate compounds were not required. The temperature range for optimum growth of cultures was 37 to 39 degrees C, and anaerobic culture conditions were essential for growth and the production of B. nodosus organisms of normal morphology. Solidified TAS media for the isolation and maintenance of B. nodosus cultures were also devised.

An agar-degrading bacterium, having a guanine-cytosine content of 50-5 mol% has been isolated from sewage. This Gram-negative rod grew well in a simple salts medium containing various carbohydrates. Growing bacteria dissolved gels and suspensions of agar and agarose rapidly, but did not attack cross-linked agars. Agarase was cell-bound in exponentially growing cultures but was released into the medium at stationary phase. Both cell extracts and culture filtrates released reducing sugars from agar solutions and prevented them from gelling. Gels were not dissolved by enzyme solutions, but the turbidity and iodine-binding properties of the agar were decreased.

Choroid plexuses from adult rats were grafted to the kidneys of other rats of the same inbred strain. Almost all the grafts survived and exhibited a multiloculated pattern of choroidal papillae. Mild but progressive enlargement of the locules was probably caused by secretion of cerebrospinal fluid. Cyclophosphamide, a drug which damages the normally situated plexus, had similar but milder effects on the plexus implants. Isografts should prove useful for other studies of choroid plexus structure and function.

Replicas of the perineurium of rabbit sciatic nerve fascicles show extensive zonulae occludentes between the flattened cells of each perineuriallamella. The junctions formed by the zonulae occludentes are of the morphologically 'tight' type. Therefore, theyare likely to represent the anatomical site of the diffusion barrier previously demonstrated by other morphological and physiological methods. Neighbouring perineurial lamellae are connected to one another by numerous maculae occludentes, some of which lie on thelateral processes of the perineurial cells. Within the meshes of the maculae occludentes and close tothem, small gap junctions can be observed.

The synaptic organization of the fleshfly (Boettcherisca peregrina) ocellus has been studied by transmission electron microscropy. Three types of neuronal element are recognized in the ocellus: (1) 70-100 retinula axons (2) four thick ocellar nerve fibres and(3) several thin ocellar nerve fibres. Synaptic connections accompanied by characteristic presynaptic ribbons have been found between these three elements in five combinations. Four of these are numerous (retinula axons symapsing onto thick ocellar nerve fibres or onto thin ocellar fibres; thin ocellar nerve fibres synapsing onto thick ocellar nerve fibres or onto other thin ocellar fibres). On or two of the thin ocellar nerve fibres are mainly presynaptic elements. The remaining thin fibres are both pre-and postsynaptic. These observations suggest that the thick ocellar nerve fibres are afferent and at least one or two of the thin ocellar nerve fibres are efferent. The function of the remaining thin fibres is not known. The fifth combination is a feedback synapse from thin ocellar nerve fibres onto retinula oxons. In addition, neuro-glial synapses have been found between thin ocellar nerve fibres and glial cells. The latter two combinations are less common and may provide alternative neuronal pathways for processingocellar input.

Pleomorphic vesicles in two different classes of 'flat-vesicle-containing profiles'(F-profiles) from the superficial layers of the superior colliculus of Galago and chimpanzee were found to be disk-like in shape as revealed by tilt analysis. In both primates, presynaptic dendrites, postsynaptic F-profiles and exclusively presynaptic F-profiles cannot be distinguished on the basis of vesicle morphology. This lends to support to the notion that F-profiles originate from one type of neuron. Modifications of the basic disk shape are interpreted as manifestations of different stages in the life cycle of individual vesicles.

Teratomas are tumours which may arise spontaneously in the testis or ovary or may be induced experimentally by the implantation of young embryos into ectopic sites. In this study the nervous tissue within mouse teratomas was investigated by light and electronmicroscopy. Characteristic neuroepithelial tubules, neurons, glia and neuropil were recognized and showed no ultrastructural abnormality apart from collagenous infiltration. Synapses were frequently observed. However, other features of mature C.N.S. tissue including myelin and complex synaptic configurations were never seen, and it was not possible to recognize distinct classes of neurons, or organizations of cells and processes which characterized specific regions of the C.N.S. The limited differentiation of the nervous tissue of teratomas is discussed with reference to the normal development of C.N.A. in vivo and in vitro. The presence of synapses in the nervous tissue of teratomas is interpreted as a reflection of an intrinsic tendency of neuronal processes to form these specialized contacts even under conditions which prevent the development of certain other characteristics of nervous tissue.

When certain intracellular and extracellular localities known to be rich in protein complexes are fixed and processed for electron microscopy, they show a reticulate precipitate which represents three-dimensional framework of material that forms the wall of polygonal lacunae. This is referred to as a stereoframework. Examples of a stereoframework described her include the presynaptic dense projections, cleft substance, postsynaptic density, the cytonet, coats of coated vesicles, reticulosomes, 'microfilamentous' network of growth cones, the glycocalyx of gut microvilli, blood plasma, precipitates of the Golgi apparatus, the chromatin of nuclei and the nuclear pore complex. The stereoframeworkappears most electron-dense when it has a very close mesh, e.g. as in the case of the dense projections. The stereoframework is assumed to have no direct relationship with themolecular architecture of the protein complexes in vivo and so can be regarded as a denaturization and precipitation artifact. This being so, attempts to elucidate the substructure of the above entities simply by inspection are fruitless. Furthermore, evidence is given that stereoframework precipitation can distort or completely obliterate organelles occupying the same locality, for example this could apply to structures such as actin filaments (perhaps running into the locality marked by a dense projection), microtubules(running into the presynaptic bag), smooth ER, tenuous connections between synaptic vesicles and the presynaptic membrane, structures within the nuclear pore complex and chromosome substructures in the nucleus. Finally it is suggested that the flat shape of synaptic vesicles (at inhibitory synapses) may be a distortion effect imposed upon the synaptic vesicles not as a result of osmotic effects, but as a conformation to the shape of a stereoframework which has been precipitated from protein complexes in the vicinity ofthe synaptic vesicles.

Sensory ganglia from 9-day chick embryos were grown on collagen coated coverslips for36 h in the presence of nerve growth factor, producing a profuse neuritic outgrowth. The cultures were then incubated for varying periods in a colloidal suspension of thorium dioxide, and the pinocytotic uptake of this marker was followed by electron microscopy. Following brief exposures (3 min), most of the labelled organelles consisted of smooth surfaced vesicles and vacuoles; with longer exposures, the bulk of the marker accumulated first in cup-shaped pre-multivesticular bodies and ultimately in multivesicular bodies. The marker was also taken up into coated vesicles, dense-cored and electron lucent tubules,dense-cored vesicles and dense bodies of the multi-layered myelin body configuration. In addition, evidence suggestive of exocytosis was also obtained; views of apparent fusion of labelled multivesicular bodies with the plasmalemma involving extrusion of vesiclesand marker particles into the extracellular space were regularly encountered following long exposures.

The ultrastructure of synaptic junctions in whole brain tissue and isolated synaptic membranes has been compared. Type 1 junctions are present in the isolated membranes,readily identified by the presence of dense-staining material associated with the postsynaptic membrane, but the dense projections present at the presynaptic membrane in intact tissue are absent. Type 2 junctions are not easily recognized because of the absence of prominent junctional densities, but apposed membranes with the appearance of type 2 junctions are seen in isolated membrane preparations. Junctions without dense-staining material are also seen among SYNAPtosomes and survive the hypotonic conditions used during isolation of the membranes. It thus seems probable that both type 1 and 2 junctions are present in isolated synaptic membrane preparations. In type 1 junctions after isolation,the postsynaptic thickening and cleft substance are together seen to be composed of an array of 200 A dense-staining subunits spanning the postsynaptic unit membrane. The relationship of this structure to the ultrastructure of the cleft substance and postsynapticthickening in intact tissue is discussed.

Synaptic membranes can be treated with detergents to yield postsynaptic densities from which the lipid unit membrane has been removed. The ultrastructure of these densities is identical to THAT of the synaptic lattice observed in undigested synaptic membranes; a1000-3000 A diameter planar array of 200 A diameter dense-staining subunits. The persistence of this synaptic lattice in the absence of lipid membrane may reflect a functional role in the maintenance of defined synaptic junctional structures in intact tissue.

We have recently developed an immunoassay that can measure thyroxine rapidly and accurately in the eluate of 40 mul of dried blood spotted on filter paper at the fifth day of life. The method is completely automated and by using the samples received by the Central Laboratory of the Quebec Network for Genetic Medicine and their follow-up facilities, we are now screening every newborn in the province of Quebec for neonatal hypothyroidism. To date, from 47,000 measurements, three newborn infants with abnormally low TBG and seven hypothyroid infants have been detected. From these data we conclude that the frequency of congenital hypothyroidism is about one in 7,000 births and that our method is effective in detecting thyroid hormone abnormalities with an acceptable percentage of false positive measurements; no false negative results have occurred to our knowledge.

Focal biliary cirrhosis is an uncommon finding in infants with cystic fibrosis, but it is present in more than a fifth of surviving children and adolescents. It was found at postmortem examination in only five of 47 infants with CF younger than 3 months, in five of 32 infants from 3 to 12 months, and in 18 of 67 children older than 1 year. In infants under 3 months, excessive mucus in intrahepatic bile ducts was seen in 11 necropsies; in 15 others there were only nonspecific periportal changes. Cholestasis was found in the livers of 18 of the 26 infants. Excessive mucus in the biliary tree was occasionally associated with periportal changes and cholestasis in older infants. The periportal changes, which are regarded as nonspecific, were never found in infants more than 1 year of age.

The frequency of coronary risk factors was documented in 4,829 school children in Muscatine, Iowa, over a 14-month period of time. Serum cholesterol levels were similar for children at all ages; the mean serum cholesterol level was 182 mg/dl (SD lus or minus 29). Twenty four percent had levels larger than or equal to 200 mg/dl, 9% were larger than or equal to 220 mg/dl, 3 % were larger than or equal to 240 mg/dl, and 1% were larger than or equal to 260 mg/dl. Casual levels of serum triglyceride increased with age: the mean level was 71 mg/dl (SD plus or minus 36) at age 6 years and 108 mg/dl (SD plus or minus 45) at age 18 years. Only 15% of the children had serum triglyceride levels of 140 mg/dl or more. Blood pressure increased strikingly with age. No child between 6 and 9 years of age had blood pressures larger than or equal to 140 mm Hg systolic or larger than or equal to 90 mm Hg diastolic. In the age group 14 to 18 years, 8.9% had systolic blood pressures larger than or equal to 140 mm Hg, 12.2% had diastolic blood pressures larger than or equal to 90 mm Hg, and in 4.4% both pressures were at or above these levels. Obesity also increased through the school years. At ages 6 to 9 years, 20% had weights relative to those of the group as a whole of larger than or equal to 110%, and 5% were larger than or equal to 130%; in the 14 to 18 years age group, 25% had relative weights of larger than or equal to 110%, and 8% were larger than or equal to 130%. These data indicate that a considerable number of school-age children have risk factors which in adults are predictive of coronary heart disease.

Propionyl CoA carboxylase deficiency was found in a 7-month-old boy who presented with attacks of vomiting, anorexia, weight loss, weakness, and hypotonia. He failed to thrive and had generalized seizures. He had propionic acidemia and hyperglycinemia; these are the manifestations of the ketotic hyperglycinemia syndrome. However, ketonuria was not a consistent part of his clinical picture, and he had at least two episodes of acute overwhelming illness, the latter one fatal, in which ketones were never found in the urine. Large amounts of pyrrolidone carboxylic acid were found in body fluids.

Evaluation of 19 patients with the Williams elfin facies syndrome, in order to more completely delineate the total spectrum of the disorder, indicates that "infantile hypercalcemia, peculiar facies, supravalvular aortic stenosis" designation which was heretofore used is inappropriate. Only 32% of the patients have evidence of supravalvular aortic stenosis and not one of them has had documented hypercalcemia, including eight patients who had a serum calcium determination in the first year of life. Rather, the most consistent features are growth deficiency which is predominantly of postnatal onset, mild microcephaly with mental deficiency, and an altered pattern of facial development which includes short palpebral fissures, a stellate pattern in the iris, medial eyebrow flare, a depressed nasal bridge with anteverted nares, and thick lips. The disorder is a sporadic occurrence of unknown etiology.

Eight patients in three families had mental retardation, characteristic facies and hands, and skeletal changes; the clinical features suggested to us that they had a syndrome previously thought to represent two entities described by Lowry and associates and by Coffin and associates, respectively. New findings include skeletal, orodental, and dermatoglyphic abnormalities and histopathologic changes suggesting that the syndrome is a heritable disorder of connective tissue. Severe expression in males and transmission through mildly affected females suggest X-linked or sex-influenced autosomal dominant inheritance.

Fetal and infant hips were analyzed in an attempt to explain some of the observed complications (particularly avascular necrosis) encountered in the treatment of hip dysplasia. Gross manipulation showed a tight interlocking of the acetabular labrum into the intertrochanteric notch of the proximal femur in the commonly used treatment positions that emphaized positional extremes. Injection studies corroborated vascular occlusion at three major sites when such positions were maintained. Marked abduction appeared to be the important position to avoid during treatment.

Cord blood from 16 premature infants and 10 full-term infants, and blood from 10 healthy adults, was incubated for 30 minutes with tritiated thymidine, after which an autoradiographic study was made of spontaneously labeling cells. Apart from a varying number of erythroblasts, myelocytes, and an occasional blast cell, two main types of spontaneously labeling cells were observed: transitional cells and large lymphoid cells, previously shown to be phagocytic. Spontaneously labeling cells were 12 times more frequent in cord blood of premature and full-term infants than in the blood of adults. No transitional cells were seen in adult blood.

Seventeen constitutionally short boys were studied throughout puberty. Nine received oxandrolone (0.1 mg/kg/day). Treatment was started before onset of puberty. Eight boys served as control subjects. No significant increase in linear growth or skeletal maturation was observed in the treated group. Likewise the peak height velocity was unchanged. Pubic hair developed similarly in both groups in relation to chronologic and skeletal age. The only significant difference was a diminution in testicular volume index during treatment after bone age of 12 years and until bone age of 14 6/12 years.

Previously published data of plasma concentrations and pharmacologic effects obtained after rapid intravenous injection and during constant rate intravenous infusion of theophylline in asthmatic children were analyzed pharmacokinetically to determine the relationship between effect on pulmonary function (improvement of forced expiratory volume in the first second) and drug level. The pharmacokinetics of distribution and elimination of theophylline are well described by a linear two-compartment open model consisting of a central and a tissue compartment. There is an essentially linear correlation between the intensity of effect and the logarithm of the amount of drug in the tissue compartment. This correlation exists both with decreasing and increasing drug concentrations, i.e., after intravenous injection and during intravenous infusion of theophylline.

The administration of Na benzoate to grown and suckling Gunn rats in single doses of 7 and 35 mg/kg failed to significantly alter serum bilirubin concentrations. These doses are comparable to quantities of Na benzoate contained in injectable diazepam used therapeutically for newborn infants. Repeated doses of 7 mg/kg in the grown rat showed no effect, as well. A single dose of 100 or 200 mg/kg of Na benzoate and repeated doses of 35 mg/kg resulted in depressed serum bilirubin concentrations. The higher concentrations of Na benzoate, however, greatly exceed amounts contained in doses of diazepam recommended for clinical use in the human neonate. The data suggest that the use of injectable diazepam, in appropriate quantities, poses no hazard to the newborn infant in terms of bilirubin toxicity. The greater affinity for bilirubin of human albumin, than that of rat albumin, may further minimize the risk.

A reinvestigation of the constituents of the Osage orange (maclura pomifera) yielded, in addition to the previously reported triterpenses (lupeol, butyrospermol, and lupan-3beta,20-diol), the pigments osajin and pomiferin, and a previously unreported constituent. The structure of this new compound was investigated. On the basis of spectroscopic and chemical data, it appeared to be an epimer of lupeol and is referred to as 19alpha-H-lupeol.

In a clinical study involving 22 normal adult volunteers, the bioavailability and pharmacokinetics of prednisone tablets with varying dissolution rates were evaluated. Serum levels were measured by a radioimmunoassay for prednisolone. Absorption rate constants and serum half-lives are presented. Substantial serum levels of prednisolone were attained quite rapidly (within 0.5 hr). The observed serum levels were statistically analyzed and fitted to the one-compartment open model with first-order absorption and elimination. A qualitative correlation between the in vitro dissolution rate and the calculated initial absorption rate constants was found. However, the in vitro dissolution rates were not predictive of the overall bioavailability of the prednisone tablets tested in terms of peak concentration and area under the serum concentration-time curve.

A technique for serial sampling of whole blood, plasma, or serum from unanesthetized, unrestrained rats is described. This technique is sufficiently rapid, reliable, and independent of the need for elaborate preparation, specialized equipment, or practiced skills to appeal to a wide range of experimental and teaching situations requiring multiple sampling from many animals. A simple surgical procedure implants a one-piece jugular cannula cut from a commercial coil of silicone polymer tubing. Multiple blood samples are almost immediately available for 5 weeks or more. Plasma or serum samples are readily obtainable from each blood sample without transfer from the syringe in which it is collected. Intravenous injection through the cannula does not prejudice later sampling protocol.

A spectrophotofluorometric method is described for the assay of thioguanine. The assay involves the oxidation of thioguanine with potassium permanganate to a fluorescent product. The method is suitable for routine quality control in laboratory preparations of thioguanine.

The commonly used 4-aminoantipyrine dye formation procedure for hexachlorophene analysis in topical formulations was modified to overcome interference due to other components. Bar soaps and nonemulsion formulations are analyzed directly, employing a chloroform back-extraction stage of the dye prior to quantitation. Hexachlorophene in emulsions and liquid soaps is determined using a TLC separation prior to dye formation.

A rapid method for determining sorbitol or mannitol in pharmaceuticals by GLC using the tris-n-butyldiboronate esters is described. An internal standard unrelated to the hexitols is utilized to avoid the problems of cross-contamination introduced by the use of other hexitols.

The chemistry of the red color formed during perchloric acid titration of chlorpromazine hydrochloride in acetic acid in the presence of mercuric acetate is discussed. Addition of ascorbic acid prevents the color formation and allows titration using a crystal violet end-point. Ascorbic acid addition also sharpens the potentiometric end-point. Ascrobic acid and its oxidation product, dehydroascorbic acid, being neutral to perchloric acid, do not interfere with the titration.

A GLC method was developed for the assay of saccharin in pharmaceutical products. The procedure requires silylation with N, O-bis (trimethylsily) acetamide and subsequent chromatography on trifluoropropyl methyl silicone, using n-octacosane as an internal standard.

The design and construction of a laboratory size dryer and other accessories suitable for investigating the drying rate kinetics of granules under controlled external conditions are described. Granulations of lactose and sulfathiazole, representing water-soluble and insoluble materials, were prepared using various commonly used binders, and their drying rates were determined. The binders and diluents affected the drying rate curves for these granulations both qualitatively and quantitatively. Granules made with starch paste and gelatin solution required maximum time and energy for drying and those made with simply syrup USP required the least among the binders studied. Generally, three linear slopes were observed when the drying rate was plotted against the moisture remaining, indicating that granulation drying may be considered as occurring through three distinct phases.

A method for the separation of 3beta-hydroxysterols from other sterols is presented. The method involves precipitating 3beta-hydroxysterols as the digitonides. The digitonide is then decomposed and separated into components using chromatography on highly cross-linked lipophilic polysaccharide gels. The digitonin in the mother liquor is separated from other sterols using the same chromatographic procedure.

The effect of compressional force on the disintegration time of tablets prepared from calcium phosphate dibasic dihydrate containing various tablet disintegrants was examined. The results show that effects of compressional force on disintegration time are of two types. The first type is that of insoluble disintegrants, e.g., starch and a cation-exchange resin, where the disintegration time initially shows a dramatic decrease. After this decrease, a further increase in compressional force appears to have no effect on the disintegration time. The second type is that of soluble disintegrants, e.g., calcium sodium alginate, sodium carboxymethylcellulose, and sodium staarch glycolate, where variation in compressional force has very little effect on the disintegration time. These results are discussed in terms of the differing mechanism whereby these substances act as disintegrants.

Hypoiodite photolysis of 3beta-acetoxy-5-pragnen-20beta-olgave 3beta-acetoxy-5-pregnene-18,20-lactone in 46% yield. Lithium aluminum hydride reduction of the latter afforded 3beta, 18,20beta-trihydroxy-5-pregnen (91% yield) which, on Oppenauer oxidation, was converted to 18-hydroxyprogesterone (66%). Lead tetraacetate oxidation followed by mild saponification gave 18-hydroxydeoxycorticosterone (58%) yield).

Small mesenteric arteries free of all extraarterial tissues were obtained from anesthetized dogs and perfused in vitro with Krebs solution. Vasoconstrictor responses of these arteries to intraarterial levarterenol and epinephrine were dose related and equivalent to those of arteries surrounded by fat and other tissues. Responsiveness was stable for at least 60 min. This simple preparation is useful for the study of vasoconstrictor phenomena uncomplicated arterial tissue.

A number of indanamines substituted at the terminal amino nitrogen with various aliphatic, alicyclic, heterocyclic, and aromatic ring systems were synthesized and screened for hypoglycemic activity. None was found to possess significant activity compared to tolbutamide.

The relative solubilities of n-alkyl p-aminobenzoates in water, proplyene blycol-water mixtures, proplyene glycol, and several other pharmaceutically important solvents can be predicted on the basis of a theoretical equation. This equation relates the activity coefficient of the hydrophobic portion of the molecule to the product of its surface area and its interfacial tension [free energy per unit area of a hydrocarbon (tetradecane) against the polar or semipolar solvent of interest]. The assumptions, conclusions, and applicability of the theorectical relationship are compared to those of the Scatchard-Hildebrand approach.

The preparation of cast films of ethylcellulose containing caffeine and salicylic acid is described. These films exhibit timed release of drugs. Release rates were found to agree with both the classical first-order equation (log drug retained against time) and diffusion-controlled release models, as exemplified by Higuchi's equations (drug release linearly related to square root of time). Mathematical analysis of the data shows that the release behavior actually conforms with the diffusion-controlled model. Literature results, reported as first order, for the release of cetylpyridinium chloride and benzalkonium chloride from polyamide films were analyzed similarly and shown to be diffusion controlled. Recommendations are made for presentation and routine treatment of drug release data to avoid ambiguity and provide useful biopharmaceutical information.

The whole plant of Swertia purpurascens Wall. (Gentianaceae) has been shown to contain five tetraoxygenated and three pentaoxygenated zanthones. These are identified as 1,5,8-trihydroxy-3-methoxyxanthone, 1,3,8-trihydroxy-5-methoxyzanthone, 1-hydroxy-3,7,8-trimethoxyxanthone, 1,3,7,8-tetrahydroxyxanthone, 1,3,5,8-tetrahydroxyxanthone, and 1-hydroxy-3,4,7,8-tetramethoxyxanthone by chemical and spectral evidence. Additionally, the crude mixture of natural xanthones has been shown to include two partially emthylated pentaoxygenated xanthones as minor entities, which yield 1-hydroxy-3,4,7,8-tetramethoxyxanthone and 1-hydroxy-3,4,5,8-tetramethoxyxanthone on methylation. This is the first time that pentaoxygenated xanthones have been found in a member of the genus Swertia. 1-Hydroxy-3,4,7,8-tetramethoxyxanthone was previously known only as a synthetic compound. The total xanthones of S. purpurascens produce significant CNS stimulant actions, consistent with some therapeutic uses of the plant extract in the Indian system of medicine. The chemotaxonomic significance of the cooccurrence of various biogenetically related chemical characters in a single plant species is appraised.

Mycophenolic acid, a novel antibiotic of low toxicity containing no nitrogen atoms in its structure, induces tumor regression in several murine solid tumor assays. It has been reported in extensive structure-activity studies that chemical modifications on the antibiotic itself reduce or eliminate antitumor activity. With the objective of antitumor activity enhancement, nitrogen-containing analogs of mycophenolic acid were synthesized according to a program directed toward the ultimate synthesis of close bioisosteres of the antibiotic. Intial efforts reported here describe the terpenoid side-chain degradation of N-geranyl-2(1H)-pyridones and N-geranylglutarimides, where the terminal isopropylidene is replaced with a carboxyl group as it occurs in mycophenolic acid. The resulting nitrogen-containing analogs of the antitumor antibiotic were inactive in the l-1210 and Walker 256 tumor systems.

A novel hydrodynamic model for drug distribution, when the drug is administered intravenously in physiological systems, is presented. In addition to obtaining results of familiar multicompartment models, the theory presents fresh insights into pharmacokinetic problems. A detailed procedure is included for evaluating drug-physiological system parameters such as the elimination rate constant, volumes of distribution, and permeability properties from the experimental time course of concentration of the central compartment.

A model was developed to detect nonlinear disposition of a drug in a tissue. The model was experimentally tested relating to salicylic acid disposition in the brain. Experimental data obtained in rats are reported for doses of 25 and 40 mg/kg ip. The parameters measured for each dose were the ratio of the area under the brain concentration-time curve to the area under the plasma concentration-time curve and the ratio of the maximum brain concentration of salicylic acid to the plasma concentration at that point in time. The ratios increased with dose; furthermore, ratios calculated using plasma concentrations corrected for plasma protein binding were dose dependent. Calculations performed on literature data for salicylic acid disposition in mouse brain corroborated the results of this sutdy. The existence of a saturable transport system for the elimination of salicylic acid from the brain is supported by the data presented. The rationale necessary to apply the model to any tissue is discussed.

2-(N-Arylcarboxamide)-3-substituted ethoxyindoles were synthesized by the reaction of 2-(N-arylcarboxamide)-3-hydroxyindoles, which were obtained by the cyclization of 2-carbomethoxyphenylglycine-substituted anilides. These 2-(N-arylcarboxamide)-3-substituted ethoxyindoles were evaluated for their in vitro monoamine oxidase inhibitory ability and in vivo monamine oxidase inhibitory property as evidence by reserpine reversal response. Their anticonvulsant activity also was determined against pentylenetetrazol-induced seizures. No definite correlation could be observed between chemical structure and biological activity.

A high-pressure liquid chromatographic method, using an absorption column and sulfabenzamide as the internal standard, is proposed for the determination of sulfacetamide sodium and its principal hydrolysis product, sulfanilamide, in eye drops. It affords an average recovery of 100.9% of added sodium sulfacetamide with a relative standard deviation of 1.9%.

A GLC method for the assay of griseofulvin in bulk and dosage forms was subjected to a wide and rigorous collaborative study. An overall recovery of 99.52 plus or minus 2.33% for three samples from 19 participating laboratories was obtained. The success of this study is ascribed to the fact that strict performance requirements are specified for the operating system.

The effects of the granule size and density on the drying rate kinetics of tablet granulations were studied using lactose and sulfathiazole granules prepared with acacia mucilage and providone solution. The drying rate kinetics consisted of three distinct phases of drying when the drying rate was platted against remaining moisture content: constant rate, first falling rate, and second falling rate periods. The effect of various statistical diameters and granular density on the drying rate during the first falling rate period was analyzed, and the mechanism of moisture migration during this phase of drying was identified.

An automated high-pressure liquid chromatographic (HPLC) method for the separation and determination of aspirin, phenacetin, and caffeine in pharmaceutical dosage forms is descreibed. Separation of these compounds for quantitation is achieved on a controlled pore glass support, utilizing a mixture of acetic acid and chloroform as the mobile phase. The method is specific, accurate, and simple and provides for the quantitation of each chromatogram in a continuous fashion every 7 min. HPLC separation of other analgesics was studied on a spherical siliceous support. The feasibility of determining free salicylic acid in analgesics also was established.

The release rates of quinine sulfate from slowly eroding, timed-release tablets prepared with various amounts of a swellable gum, carbomer, and cellulose acetate hydrogen phthalate at different compaction pressures were attained. For the dissolution test of the prepared tablets, the method described in NF XIII was followed. The concentration of the released quinine sulfate was determined spectrophotometrically.

The first-pass equation based on the dose, hepatic blood flow, and total area under the plasma level-time curve after oral administration was used retrospectively to predict the bioavailability of nortriptyline after oral administration of 1 mg/kg-single oral doses to monozygotic and dizygotic twin pairs. The predicted values of bioavailability ranged from 45 to 85%, consistent with experimentally derived estimates of nortriptyline availability.

The effects of acids and bases on the viscosity of salicylic acid-cetrimide systems were investigated. A viscosity reduction was produced by the addition of acids and was independent of the degree of saturation of the cetrimide solution with salicylic acid. The incorporation of base followed by that of acid increased and decreased the viscosity, respectively. This viscosity behavior was demonstrated in undersaturated systems and was not goverened by the relative amounts of base or acid used. In oversaturated systems, only a lowering of viscosity was observed.

Aliphatic aldehydes react with 2,3-dimethyl-2,3-bis-(hydroxylamino)butane and sodium periodate for form colored free radicals. These radicals were stabilized with pyridine in aqueous solution. Low levels of formaldehyde in aqueous solutions were determined utilizing this reaction.

A sensitive procedure was developed for metronidazole in human plasma. The metronidazole is extracted from the plasma with chloroform and determined as the trimethylsilyl derivatives by GLC using a flame-ionization detector. Myristyl alcohol is used as the internal standard for quantitation by relative peak height.

A study was undertaken to develop a method that could be used an an indication of the absorption of steroids. It was demonstrated that 17-hydroxycorticosteroids in urine could be quantitatively determined via absorptivity values obtained through the use of standard hydrocortisone-alcohol solutions or standard solutions of hydrocortisone in urine utilizing the blue tetrazolium assay method. This method is dependent upon the hydrolysis of conjugated urinary steroids by beef liver glucuronidase. The resultant concentration of 17-hydroxycorticosteroids is determined colorimetrically using blue tetrazolium.

Binding of trichloromonofluoromethane, dichlorodifluoromethane, and dichlorotetrafluoroethane was studied in aqueous 5% human albumin solution, using the partition coefficient method in sealed serum bottles. The partition coefficient and the fraction of fluorocarbons bound were highly dependent on fluorocarbon concentrations. The average binding sites per molecule of albumin were 2.17, 0.30, and 0.42 and the binding association constants were 1.11 X10-3, 1.73 X10-3, and 5.06X10-3 M-minus 1, respectively. At the lowest concentration studied, 62.3, 25.5, and 65.6% were found bound to albumin, respectively. This appears to represent the first extensive study on any gas-albumin interaction.

The release of sulfathiazole and salicylic acid from white petrolatum containing ethoxylated surfactants of varying ethylene oxide chain length was evaluated by the dialysis method. The rate of release of the drugs increased with the ethylene oxide number of the added surfactant.

Treatment of mixed anhydrides derived from cis- and trans-2-carbobenzyloxycyclopropanecarboxylic acids and ethyl chloroformate with ethoxymagnesio di-tert-butyl malonate and subsequent treatment of the resulting adducts with p-toluenesulfonic acid afforded cis- and trans-benzyl-2-acetylcyclopropanecarboxylates in good to excellent yields, with retention of the original stereochemistry of the systems. These methyl ketones and an open chain congener, benzyl levulinate, were inert toward m-chloroperbenzoic acid. The cis-isomer and benzyl levulinate underwent normal Baeyer-Villiger reactions mediated by trifluoroperacetic acid, forming moderate yields of the acetate ester insertion products.

The syntheses of 2-(9-acridinyl)ethyl-N-substituted carbamates and their hydrochlorides and 10-N-oxides are reported along with biological results in the areas of antineoplastic, antimalarial, and CNS activity screening. The compounds showed negative biological activity in the areas tested.

A comparison of the bioavailability of salicylate from five brands of commercially available aspirin rectal suppositories in an adult panel is presented. All brands show slow absorption compared to oral administration of the drug in tablet form. At best, about 40% of the dose (on the average) was absorbed when retention time in the bowel was limited to 2 hr. However, four out of the five brands give substantially lower absorption rates so that only about 20% of the aspirin is available.

The activity of tertiary and quaternary anticholinergic drugs was compared in two different test procedures designed to measure cholinolytic activity in mice. The four drugs utilized were atropine sulfate, atropine methylnitrate, scopolamine hydrobromide, and scopolamine methylnitrate. The results led to the conclusion that one of these test procedures, the induction of mydriasis (increase in pupil size), primarily measures peripheral anticholinergic activity whereas the other procedure, inhibition of physostigmine lethality, primarily measures anticholinergic activity in the CNS. These two test procedures can be utilized to characterize the nature of the cholinolytic properties of prospective therapeutic drug candidates.

The distribution of radioactivity as 5-, 10-, and 15-min intervals following the intravenous administration of N-14-C-azure C was determined. The concentrations of radioactivity observed indicated that radioactive derivatives of azure C would not be useful pancreas of parathyroid scanning agents.

The influences of drug concentration and vehicle composition on the corneal penetration of the steroid fluorometholone were studied in the albino rabbit. Aqueous dosing systems included a saturated solution and 0.1, 0.05, and 0.1% suspensions of micronized fluorometholone. Two different doses of a 0.1% oleaginous ointment were also studied. The results from the 0.1 and 0.5% suspensions show a peak aqueous humor steroid concentration at 30 min and a substantial sustaining effect with these two concentrations. The results also support the belief that moderate dilution of a suspension of a slowly soluble drug may not substantially lower the aqueous humor drug levels or, conversely, that use of a higher concentration suspension may not improve the aqueous humor drug concentration-time profile. The 0.1% suspension and the saturated solution did not produce a sustaining effect. The results demonstrate for the first time that the particles present in a dose of suspension are retained within the cul-de-sac of the eye and contribute significantly to the amount of steroid penetrating the cornea. This finding was confirmed by a study in which the eye was rinsed with saline solution 30 min after instillation of a dose of a 0.05% suspension. The rinsing procedure prematurely terminated the sustaining effect of the suspension. The results of the ointment studies show that partitioning of the lipophilic steroid from the oleaginous vehicle has a greater rate-limiting influence on corneal penetration than the dissolution rate parameter associated with the aqueous suspensions. Peak aqueous humor concentration was not achieved until 3 hr after dosing and was comparable to the 0.1 and 0.05% suspensions. Predosing of the eye with a saturated solution or the 0.1% suspension prior to dosing with ointment overcomes the inability of the ointment to provide adequate drug at short times following dosing. In this case, peak levels were achieved within 60 min and then maintained. The duration of aqueous humor levels and the amount penetrating from the ointment were greater than the suspensions, and these effects are discussed relative to the mechanism. Differences in aqueous humor levels produced by 25- and 50-mg doses of ointment were minimal. A discussion of the results from all studies is presented in the context of present theories regarding the role of the lipophilic epithelial layer of the cornea as a barrier to drug penetration.

A simple, rapid, and sensitive radioimmunoassay was developed for the determination of chlordiazepoxide (7-chloro-2-methylamino-5-phenyl-3H-1,4-benzodiazepine 4-oxide) in plasma. Antiserum capable of binding chlordiazepoxide-2-14C was obtained from rabbits following immunization with an antigen prepared by coupling the reactive acyl azide of 7-chloro-5-(4-hydrazinocarbonylmethoxyphenyl)-2-methylamino-3H-1,4-benzodiazepine 4-oxide to bovine serum albumin. The radioimmunoassay allows for the specific determination of chlordiazepoxide directly in plasma without extraction and was compared with a differential spectrofluorometric assay for chlordiazepoxide. Both methods gave satisfactory agreement for the plasma levels of chlordiazepoxide in human subjects resulting from single and chronic oral doses of chlordiazepoxide hydrochloride.

Aluminum hydroxide gel loses reactivity upon aging. However, a sharp decrease in reactivity occurs when the gel is diluted with double-distilled water. The loss of reactivity is directly related to the degree of dilution, but dilution with dioxane or the mother liquor has no effect on reactivity. It is hypothesized that dilution with water causes a change in the equilibrium between stabilizing ions incorporated in the gel structure and ions in solution. As the stabilizing ions leave the gel structure to reestablish the equilibrium, a loss of reactivity is observed until a new equilibrium is established.

The applicability of a permeation rate technique to the determination of drug-macromolecule interactions was tested by measuring the extent of interaction of methylparaben with polyvinylpyrrolidone and polysorbate 80. Results were in agreement with literature data obtained by other techniques. The present method, although restricted to permeant molecules that diffuse readily through nonporous nylon membranes, is of potential value for investigations of drug binding by macromolecules not retained by porous dialysis membranes.

Reaction of chlorpromazine with 9-bromomethylacridine under appropriate conditions yields a nonfluorescent quaternary ammonium derivative which, on subsequent photolysis, liberates fluorescence. The major component of this fluorescence is 9-methylacridine (86%), while two minor components are 9-acridinecarboxaldehyde (6%) and 9-acridinemethanol (8%). The mechanism of photolysis leading to formation of these products appears to involve homolytic as well as heterolytic cleavages of the quaternary salt. Both the quaternization and the photolysis are stoichiometric. Appropriate isolation of the fluorescence and its quantitative determination constitutes the basis of a new and highly sensitive assay applicable to chlorpromazine and other tertiary amine drugs.

The 3, 3-dimethyl-, 3-n-butyl-3-methyl-, 3-(2-hydroxyethyl)-3-methyl-, 3,3-bis(2-fluoroethyl)-, and 3, 3-bis(2-chloroethyl)-1-triazenyl derivatives of imidazole-4-carbonitrile were prepared from 5-diazoimidazole-4-carbonitrile, a stable compound which produced a mass spectrum consistent with its structure. In contrast to the corresponding carboxamides, none of the triazenylimidazole-4-carbonitriles were active against lymphatic leukemia L-1210 im mice. Like the analogous carboxamide, the bis(2-chloroethyl) triazene readily cyclizes to the 1, 2, 3-triazolinium chloride.

The possibility tht metronidazole exerts several of its biological actions via interaction with important metal ions was investigated. NMR spectroscopy and polarography (ac) were used to test for any interaction, to locate the probable sites for complexation, and to determine the molecular stoichiometry of any complexes formed. Of the series of divalent metal ions tests, only cupric ion showed detectable interaction with metronidazole. The predominant site of interaction of cupric ion was the unsubstituted nitrogen atom (N-3) on the metronidazole molecule. The stoichiometry of the complex was (Cu-(metronidazole)4)+2. A likely structure for the complex is presented.

The extent of plasma binding and the partition coefficient of disopyramide and 20 disopyramide derivatives were determined. Structural variations on the four functional groups around the tetrahedral carbon in the disopyramide molecule was found to influence both parameters to varying degrees. Three linear equations were developed to correlate the observed effects, depending on the type of chemical modification. The linear correlation between drug-plasma interaction and lipophilic character was analyzed theoretically. A simple model was derived to relate quantitatively the variation in the extent of plasma binding to the change in lipophilicity of disopyramide derivatives.

The role of principal component analysis in the selection of pharmaceutical formulations is presented. The objective and the procedure of the analysis are discussed in detail. The technique was successfully applied to a system consisting of 10 response variables (tablet properties). Analysis of the results showed that the first component (dissolution) and components one and two together (dissolution and disintegration) contributed 95.4 and 99.3%, respectively, to the overall information about the formulations and that eight of 10 response parameters contributed nothing further to the overall information. The results obtained from this method of analysis may be found useful for achieving economy in both cost and time of measuring response. Principal component analysis also provides a basis for understanding the underlying mechanism of the system under consideration.

The published procedure for the synthesis of pyrrolidone-5-hydroxamic acid was improved. The acidity constant of the pyrrolidone-5-hydroxamic acid was determined as pKa = 8.65. In an aqueous solution of iron (III) ions, pyrrolidone-5-hydroxamic acid binds ferric ion, forming a mixture of mono-, bis-, and tris(pyrrolidone-5-hydroxamato)iron (III) complexes. These complexes were studied by potentiometric and spectrophotometric methods. The tris compound was isolated as dark orange-red crystals and identified according to elemental analysis and IR spectral data as C15H21FeN6O9.6H2O, having the magnetic moment of 5.67 B.M.

A spectrodensitometric method for the direct determination of sulfadiazine at the tissue residue level (0.1 ppm) is based upon the measurement of the fluorescence of a sulfadiazine-fluorescamine derivative formed directly on a TLC plate by dipping it into a fluorescamine solution. The linear dynamic range for the assay is about 150 from 200 to 0.2 ng, the lower limit of sensitivity. Recoveries from various spiked tissues including milk, eggs, liver, kidneys, muscle, skin, and fat varied with the tissue type but were reproducible. The assay technique has also been used for the assay of sulfamethoxazole and has been explored for use in specifically assaying sulfonamide mixtures.

Subjective spreadability, viscosity, and stickiness perceived with the fingers were predicted from fluid mechanics. The correlation coefficients of these predictions were 0.95 for spreadability, 0.95 for viscosity, and 0.09 for stickiness. The two important assumptions in the predictions were that spreadability and viscosity were perceived as shear stress and that stickiness was perceived as time. When the finger geometry was approximated as two parallel plates, the predictions only required rheological data for the Newtonian and non-Newtonian liquids used. While these liquids covered a range of 10-6 in apparent viscosity, large variations in other fluid properties such as density and surface tension were not studied.

The shifts in the absorption and fluorescence spectra of 3-aminoacridine, proflavine, acridine orange, and acridine yellow were employed to show that the singly charged cations, the predominant species at biological pH, exist in the ground state in the amino form. In the lowest excited singlet state, however, the monocations of the diaminoacridines have the imino structure, a conclusion supported by the relative ground- and excited-state pKa values of the reactions of the monocation with H-+. The ground-state amino structure has its positive charge concentrated at the heterocyclic nitrogen atom, a fact that is of primary importance in determining the geometry of binding to DNA.

The influence of dioctyl sodium sulfosuccinate and poloxalene on the GI absorption of phenolsulfonphthalein in the rat was studied. Urinary excretion data after oral administration of the drug to intact rats and loss of the drug from the whole small intestine as a loop were both utilized to assess the effect of the surfactants on absorption. Dioctyl sodium sulfosuccinate markedly increased the absorption of the drug, and the extent was dependent on the surfactant concentration. Maximum effect was observed at the reported ED50 in rats, of the surfactant as a fecal softener. The mechanism responsible for absorption enhancement seems to be an alteration of the permeability of the intestinal membrane. Micellar complexation between the drug and the surfactant resulted in a lesser increase in absorption at the higher surfactant concentrations. Poloxalene did not increase drug absorption, but higher concentrations caused a decrease in absorption due to micellar entrapments of the drug molecules. The influence of dioctyl sodium sulfosuccinate on the peritoneal absorption of the drug was also investigated. Lower doses of the surfactant increased absorption of the drug by altering the membrane permeability. Higher doses decreased absorption due to unavailability of the drug molecules entrapped in the micelles.

To explore the effect of dioctyl sodium sulfosuccinate on drug absorption in humans, the urinary excretion of a poorly absorbable drug, phenolsulfonphthalein, administered in solution with and without the surfactant was determined. Coadministration of a therapeutic dose of the surfactant with the drug solution resulted in a significant increase in the initial rate of absorption. A small increase in the extent of absorption was also observed. Pretreatment with the surfactant for 6 nights, followed by administration of the drug on the 7th day, did not significantly change the rate of extent of absorption. The surfactant is thought to have a direct effect on the GI membrane, resulting in a temporary change in its permeability. This effect appears to be reversible after a few hours.

Specific assay procedures were developed to measure plasma concentrations of (plus or minus)-1, 2-bis(3, 5-dioxopiperazinyl) propane (1) by GLC using flame-ionization detection with a sensitivity limit of 5 mug/ml and by GLC-mass fragmentography with a sensitivity limit of 0.2 mug/ml. Applicability of the assay procedures was demonstrated in rats, rabbits, and humans. Plasma concentration-time curves of total 14-C activity and intact I was obtained in rats and rabbits following oral and intravenous administration of 14-C-I. Plasma elimination half-lives of I in the first 2 hr following intravenous doses in rats were 40 and 45 min in two rats. Plasma levels of I were measured over 6 hr after an intravenous dose in rabbits and followed a two-compartment open model with a terminal loglinear plasma half-life of 85 min. Significantly higher total 14-C levels compared to intact I plasma concentrations indicated rapid biotransformation in both rats and rabbits to unknown metabolites. The oral bioavailability appeared to be limited in both species relative to intravenous administration. Two patients receiving 3 g I/m-2 in tablets orally showed plasma levels of I similar to those obtained after oral doses in rats and rabbits, with peak concentrations at 2 hr after the dose (3.8 mug/ml) and with still measurable levels 12 hr after the dose (0.4 mug/ml).

At 37 degrees C the vasoconstrictor response of the isolated, perfused rabbit ear artery to an infusion of nicotine or acetylcholine (ACh) was transient--rising rapidly to a peak and then fading completely within a minute or two. After complete fade of the response during continued infusion of either nicotine or ACh (in the presence of atropine), the vasoconstrictor responses to stimulation of periarterial adrenergic neurons or to infused norepinephrine (NE) were not diminished. The fade of response was attributed to desensitization of nicotinic receptors at the adrenergic nerve terminals on which nicotine and ACh act to release NE. On washout of either nicotinic agonist after development of desensitization, recovery of sensitivity was essentially complete within several minutes, provided that neither excessively high concentrations nor excessively long periods of infusion had been used. The rate of desensitization to nicotine of ACh increased with concentration of the agonist. On infusion of high concentrations (200 mug/ml) of either, the time required for full desensitization was estimated to be less than 5 seconds. Cross-desensitization was demonstrated for nicotine, ACh and tetramethylammonium. Considerable desensitization occurred even on infusion of nicotine (1 mug/ml) slightly below that required to give a threshold vasoconstrictor response. Moreover, full desensitization without any preceding vasoconstrictor response could be obtained if the concentration of infused nicotine was gradually increased from an initial subthreshold to a final very high suprathreshold level over a 20- to 30-minute period. Both the rate and degree of desensitization to nicotine decreased as the temperature was decreased from 37 degrees to 27 degrees C. The present results are consistent with the concept that the nicotinic receptor (or receptor mechanism) at the adrenergic nerve terminal, after being activated as a result of combination with the agonist, can undergo a transformation to an inactive or desensitized state. It is proposed that under conditions where desensitization to the agonist develops in the absence of any vasoconstrictor response, the fraction of receptors in the activated state at any instant in time during the development is too small to trigger the release of NE.

Isomers of amphetamine were tested for their ability to evoke changes in the spontaneous motor behavior of rats. d-Amphetamine was effective within the dose range of 0.2 to 2.0 mg/kg; l-amphetamine was effective within the range of 1.0 to 10.0 mg/kg. Both d- and l-amphetamine were also compared for their pressor and tachycardic activity in pithed rats. The doses of amphetamine tested (0.1-10.0 mg/kg) were identical to those which produced changes in behavior. d-Amphetamine was approximately 5-fold more potent than l-amphetamine in evoking pressor responses and approximately 3-fold more potent in evoking tachycardic responses. The two isomers differed little in their ability to block norepinephrine uptake into heart. The ID50 values were; d-amphetamine 0.7 mg/kg; l-amphetamine, 1.2 mg/kg. Cocaine, which also blocked norepinephrine uptake into heart (ID50 = 3.0 mg/kg), lacked significant pressor or tachycardic activity in pithed, adrenalectomized animals. The inability of cocaine to evoke responses was related to the marked reduction in norepinephrine turnover and release in pithed animals. It was concluded that differences in potency between d- and l-amphetamine cannot be explained on the basis of blockade of norepinephrine uptake. The difference may relate to evoked release of amine.

The effects of chronic ingestion of oral contraceptive preparations with "low" and "high" estrogen contents on conversion of angiotensin I (AI) to angiotensin II (AII) have been examined in the rat. The dosage employed was 1.0 mg/kg/day for a period of 3 to 4 weeks. Blood pressure responses in anesthesized animals showed that in vivo conversion of AI was increased by about 40% with either drug treatment. The activity of injected renin was increased to the same extent as that of AI following treatment with the low-estrogen preparation, and to a greater extent following treatment with the high-estrogen preparation. Blood pressure responses to AII were also slightly increased above control values in those animals which received high-estrogen treatment. However, conversion of AI in isolated perfused lungs or in plasma from treated rats was not significantly altered from normal. It is concluded that conversion of AI to AII can constitute the rate-limiting step in AII production under certain circumstances and that increased conversion may play a part in the changes in the renin-angiotensin system associated with high circulating estrogen levels. The increased conversion is unlikely to be due to a change in pulmonary or plasma converting enzyme activity.

The antiarrhythmic efficacy of ajmaline has been evaluated in three experimental models of cardiac arrhythmias in the dog. These data have been related to the actions of the compound on several parameters of heart function and compared to results obtained in various clinical arrhythmias. Ajmaline was more effective in arrhythmias of the ectopic focus type than in the circus movement model. These results agreed with the pattern of clinical activity. The compound produced decreases in excitability and conduction and increased the functional refractory period in all heart tissues. These effects were most marked in the atrium. The drug also showed a moderate degree of anticholinergic activity. Both in the dogs and in the clinical cases, the agent showed an important hypotensive effect. In a group of experiments in which transmembrane potentials were recorded, the compound produced in all tissues a decrease in upstroke velocity and amplitude of the action potential; it also increased the duration of the action potential in atrial and ventricular muscle, but it decreased it in Purkinje fibers. The possible mechanism(s) of action of the drug is discussed in terms of the different hypotheses for cardiac arrhythmias.

Bretylium produced electrophysiological effects on both rat atrium and ventricle in vitro at concentrations ranging from 2 times 10- minus 5 to 10- minus three M. Those effects included lengthening of action potential duration and effective refractory period; increasing effective refractory period/action potential duration; decreasing dv/dt of phase zero of the action potential and suppressing the action potential amplitude and overshoot. These effects, which could serve as a basis for the antiarrhythmic action of bretylium, were observed also in hearts from immunosympathectomized rats confirming a direct effect of this drug on the electrical properties of the cardiac muscle cells. In vivo and in vitro exposure of the myocardium to 14-C-bretylium showed that this drug is concentrated in cardiac ventricle and that this concentrating ability of the heart may be responsible for attaining effective antiarrhythmic concentrations in the myocardium at low plasma concentrations of the drug. Uptake of bretylium by the sympathetic nerves never amounted to more than 15% of the total bretylium binding by the cardiac ventricle and this neuronal uptake became insignificant compared to total bretylium uptake at concentrations greater than 10- minus 6 M. Subcellular distribution of the bretylium bound to the cardiac ventricle from immunosympathectomized rats suggested a binding to plasma membranes. Efflux studies indicate that this binding was tight, although reversible. These results indicate that underlying the antiarrhythmic effects of bretylium is an accumulation of the drug by cardiac muscle cells and a direct effect of the drug on the electrical properties of the cardiac muscle membrane independent of any action on the adrenergic neuron.

The effects of dimethyl quaternary propranolol (UM-272) on electrophysiologic properties of canine cardiac Purkinje fibers (PF) were studied using standard microelectrode techniques. In PF superfused with Tyrode's solution, the effects of UM-272, 10- minus 7 to 10- minus 5 M, were studied. In other experiments, UM-272, 3 mg/kg, was injected into donor animals whose blood was used to superfuse isolated PF. Antiarrhythmic concentrations of UM-272 decreased action potential (AP) amplitude, phase 0 upstroke velocity, membrane responsiveness, AP duration and the effective refractory period (the decrease in the effective refractory period was less than that in AP duration). Automaticity of spontaneously firing PF was suppressed. As perfusate potassium concentration was increased, the magnitude of UM-272 effect was accentuated. When the actions of equimolar concentrations of UM-272 and propranolol on PF AP were compared, propranolol attained a steady-state effect more rapidly and tended to depress the AP more markedly. UM-272 did not block epinephrine-induced increases in PF automaticity or the adenylate cyclase activation of PG homogenates induced by epinephrine. These studies indicate that UM-272 has direct effects on the PF AP similar to those of propranolol, but lacks the beta blocking properties of the latter.