CO2 capture by means of an enzyme-based reactor.

We report a means for efficient and selective extraction of carbon dioxide (CO(2)) at low to medium concentration from mixed gas streams. CO(2) capture was accomplished by use of a novel enzyme-based, facilitated transport contained liquid membrane (EBCLM) reactor. The parametric studies we report explore both structural and operational parameters of this design. The structural parameters include carbonic anhydrase (CA) concentration, buffer concentration and pH, and liquid membrane thickness. The operational parameters are temperature, humidity of the inlet gas stream, and CO(2) concentration in the feed stream. The data show that this system effectively captures CO(2) over the range 400 ppm to at least 100,000 ppm, at or around ambient temperature and pressure. In a single pass across this homogeneous catalyst design, given a feed of 0.1% CO(2), the selectivity of CO(2) versus N(2) is 1,090 : 1 and CO(2) versus O(2) is 790 :1. CO(2) permeance is 4.71 x 10(-8) molm(-2) Pa(-1) sec(-1). The CLM design results in a system that is very stable even in the presence of dry feed and sweep gases.

Carbon dioxide transport by proteic and facilitated transport membranes.

Membrane separation of gases is governed by the permeability of each species across the membrane. The ratio of permeabilities yields the selectivity. Use of certain organic carriers in facilitated transport membranes and the CO2 converting enzyme carbonic anhydrase (CA) in proteic and facilitated transport membranes allows a dramatic increase in CO2 selectivity over other gases. CA has a low Km (9 mM), which we predicted would allow it to scavenge CO2 to very low partial pressures. Our goal was to determine if CA could remove CO2 from an environment at levels of 0.1% or less. Prior measurements of CO2 transport across thin supported liquid membranes showed that addition of CA enhanced CO2 flux by 3- to 100-fold. Proteic films use bifunctional reagents (e.g., glutaraldehyde) to cross-link the enzyme forming a gel. Bovine serum albumin (BSA) is often added for structural stability. Using such a preparation we examined the ability of proteic films to improve CO2 selectivity and to scavenge CO2 from a mixed gas stream. Proof-of-concept results, measured by mass spectrometry, showed a fivefold improvement in CO2 capture rate with maximal improvement at CO2 values of 1% partial pressure difference in the presence of 0 atm absolute difference. At 0.1% CO2 the membrane exhibited a 76% improvement over controls. At 0.3% CO2 the improvement is about threefold. CA proteic membranes exhibit selectivity for CO2 over oxygen and nitrogen in excess of three orders of magnitude. A CA-based proteic or facilitated transport membrane should readily achieve CO2 partial pressures of 0.05% under CELSS conditions. In addition to proteic membranes we are exploring direct immobilization of engineered CA to ultra-high-permeability teflon membranes. Site-directed mutagenesis was used to add functional groups while retaining full enzymatic activity. These results provide a basis for development of far more efficient CO2 capture proteic and facilitated transport membranes with increased selectivity to values closer to 100-fold at 1% CO2. The result will be CO2 selectivity at 0.1% on the order of 400-fold. These results exceed those obtained with other technologies.

Enhancement of attention processing by Kantroll in healthy humans: a pilot study.

This is the first report in humans of the effects of daily ingestion of a specific amino acid mixture, Kantroll, on cognitive event-related potentials (ERPs) associated with performance. Cognitive ERPs were generated by two computerized visual attention tasks, the Spatial Orientation Task (SOT) and Contingent Continuous Performance Task (CCPT), in normal young adult volunteers, where each subject acted as his own control for testing before and after 28-30 days of amino acid ingestion. A statistically significant amplitude enhancement of the P300 component of the ERPs was seen after Kantroll for both tasks, as well as improvement with respect to cognitive processing speeds. The enhancement of neurophysiologic function observed in this study on normal controls is consistent with the facilitation of recovery of individuals with RDS (i.e., substance use disorder, ADHD, carbohydrate bingeing) following the ingestion of the amino acid supplement, Kantroll, and warrants additional placebo-controlled, double-blind, studies to confirm and extend these results.

Neurodynamics of relapse prevention: a neuronutrient approach to outpatient DUI offenders.

The central nervous system rewarding properties of ethanol, cocaine, and heroin may activate a common catecholaminergic reward system in the mesolimbic circuitry of the brain. Driving-under-the-influence (DUI) offenders with either alcohol- or cocaine-related problems were studied. The neuronutrients SAAVE and Tropamine significantly reduced relapse rates and enhanced recovery in these DUI outpatient offenders over a 10-week period. Follow-up on both the SAAVE and Tropamine groups after 10 months revealed a 73% and a 53% overall recovery rate, respectively. These clinical results favor the use of these neuronutrients as adjuncts to psychological therapeutic modalities.

Ethanol ingestive behavior as a function of central neurotransmission.

Uncontrollable alcohol ingestive behavior has been linked to deficits of central neurotransmission. The pineal gland plays an important role in modulating ethanol intake in numerous animal species. The opioidergic (i.e. beta-endorphin, enkephalin, and dynorphin) system is involved in both the actions of alcohol and opiates, as well as craving and/or genetic predisposition towards abuse of these two agents. Furthermore, there is significant evidence to link ingestive behaviors with the ventral tegmental accumbens-hypothalamic axis, whereby the biogenic amines dopamine and serotonin are reciprocally involved. Evidence is presented which implicates the striatum and the hypothalamus as possible specific loci for regional differences between alcohol-preferring and alcohol-nonpreferring mice. We believe that photoperiod-induced alcohol ingestive behavior may involve alterations in both pineal and hypothalamic opioid peptides.

Enkephalinase inhibition and precursor amino acid loading improves inpatient treatment of alcohol and polydrug abusers: double-blind placebo-controlled study of the nutritional adjunct SAAVE.

We investigated the effects of the amino acid and vitamin mixture SAAVE in inpatient, chemically-dependent subjects to evaluate the role of neurotransmitters in facilitating recovery and adjustment to a detoxified, sober state. SAAVE is formulated from amino acids that are precursors for neurotransmitters and neuromodulators thought to be involved in alcohol and drug seeking behavior. In a double-blind, placebo-controlled, randomized study of 62 alcoholics and polydrug abusers, SAAVE patients had a significantly reduced stress response as measured by the skin conductance level (SCL), and significantly improved Physical Scores and BESS Scores (behavioral, emotional, social and spiritual). After detoxification there was a six-fold decrease in AMA rates when comparing SAAVE vs. placebo groups. In this inpatient treatment experience SAAVE facilitated the rate of recovery and allowed patients to respond more fully and more quickly to the behavioral goals of the program, for example as measured by the BESS Score. The use of SAAVE to achieve enkephalinase inhibition and precursor amino acid loading in the acute inpatient treatment environment provides the practitioner with the potential ability to restore the neurochemical changes inherent to alcoholism and drug abuse. These findings increase our understanding of the clinically relevant neurobiological mechanisms which underlie compulsive disease.

Improvement of inpatient treatment of the alcoholic as a function of neurotransmitter restoration: a pilot study.

We report results of a double-blind evaluation of the nutritional supplement SAAVE for facilitating improvement in a 30-day inpatient alcohol and drug rehabilitation center. SAAVE is uniquely designed to elevate levels of enkephalin(s), serotonin, catecholamines, and GABA, which are believed to be functionally deficient in alcoholics. Twenty-two patients were studied. The SAAVE patients, as compared to the control group (a) had a lower BUD (building up to drink) score, 1 vs 2; (b) required no PRN benzodiazepines, 0% vs 94%; (c) ceased tremoring at 72 h, as compared to 96 h; and (d) had no severe depression on the MMPI, in contrast to 24% of control group. These preliminary data suggest that SAAVE is a valuable adjunct to therapy by aiding the patient's physical adjustment to a detoxified state while facilitating a more positive response to behavioral therapy.

Alcoholism: scientific basis of a neuropsychogenetic disease.

Until recently alcoholism was regarded as being an incurable psychological problem. During the last decade a chain of research has led to a new insight into the causes and potential alleviation of alcohol craving: Recent discoveries indicate that the brain has receptor sites for naturally occurring opiate-like substances (endorphins and enkephalins) which are produced by the nervous system. Opiates such as morphine or heroin, and some of the metabolic products of alcohol (tetrahydroisoquinolines), can also attach themselves to these receptors. It has been further discovered that the craving for alcohol is related to a deficiency of the naturally occurring opiate-like substances as well as other neurotransmitter substances. This deficiency can occur genetically or as a result of prolonged stress or long-term heavy drinking. The neurochemical imbalance may be treated chemically, leading to a possible alleviation of the craving for alcohol, especially in conjunction with psychotherapeutic and counseling regimens.

Enkephalinase inhibition: regulation of ethanol intake in genetically predisposed mice.

This is the first report of alteration in alcohol intake in mice with a genetic predisposition to alcohol preference and known to have innate brain enkephalin deficiencies. We have been able to significantly attenuate both volitional and forced ethanol intake respectively by acute and chronic treatment with hydrocinnamic acid and D-phenylalanine, known carboxypeptidase (enkephalinase) inhibitors. Since these agents, through their enkephalinase inhibitory activity, raise brain enkephalin levels, we propose that excessive alcohol intake can be regulated by alteration of endogenous brain opioid peptides.

Alcohol and opioid peptides: neuropharmacological rationale for physical craving of alcohol.

Until recently alcoholism was regarded as an incurable psychological problem. During the last decade a chain of research has led to important hypotheses about the etiology of the physical craving of alcohol. Recent discoveries indicate that the brain has receptor sites for naturally occurring opiatelike substances (endorphins, enkephalins, and dynorphins) which are produced by the nervous system. Opiates such as morphine or heroin and some of the metabolic products of alcohol (tetrahydroisoquinolines) can also attach themselves to these receptors. It has been further discovered that the physiological craving for alcohol may be the result of a deficiency of the naturally occurring opiatelike substances as well as other neurochemical deficits (i.e., dopaminergic, GABAergic, and serotonergic). These neurochemical deficits can occur genetically or as a result of long-term heavy drinking.

Primary retinal and cortical glial cell cultures: effects of medium and serum on attachment and growth.

Glial cells of the retina are anatomically distinctive and are thought to contribute importantly to retinal electrophysiology. However, no adequate preparation exists for studying them in isolation, in vitro. This report provides guidelines for primary retinal glial cultures (RET) and compares basal tissue culture features with those for neocortical glia (CX) and the well-studied rat glial line, C6. Cell attachment and growth of RET, CX, and C6 are unique. These differences are explored by the use of specific media and sera. RET attachment, unlike that for CX or C6, was far more sensitive to medium than serum. RET cells attached least quickly, CX most quickly; 4 hr after plating 20% of RET remained unattached. RET growth was poor and relatively insensitive to medium. In contrast, growth of CX or C6 was medium dependent. Serum had substantial effects on the growth of all three glial lines. Pig, goat, horse, and dog sera were particularly effective, often comparing favorably to fetal calf serum. Medium or serum optimal for cell attachment, typically, was not optimal for growth and serum effects were more dramatic than those of medium. By all measures, CX and C6, both derived from brain, were more alike than were the two rabbit primaries, CX and RET. The data reveal substantial differences between presumably similar cells and indicate a need for an empirically based choice of both basal-salt media and serum to optimize specific aspects of cell development in culture.

Potential use of bar codes to implement automated dispensing quality assurance programs.

Bar code-based systems have automated many counting, tracking, and sorting functions in health care delivery services. We designed and briefly tested the feasibility of a bar code-based dispensing quality assurance system for a hospital outpatient pharmacy. The use of bar codes to verify the identity of the dispensed product required an extra few seconds processing time for each prescription but did not increase markedly the processing time when compared to a control period. In addition to verifying product identity, the system checked the manufacture's expiration date to reduce the risk of delivering outdated medications to the patient. The on-site test period for this feasibility model was relatively short (one week) and no actual dispensing errors were detected. However, when the system was presented with 100 different prescription forms containing 50 randomly sequenced, precontrived dispensing errors, the system identified all medication errors and outdated products. As shown in other studies, bar code-based systems have the potential to capture information not effectively recorded using manual methods. We suggest they should be considered by pharmacists interested in automating inventory management and work flow functions or establishing automated dispensing quality assurance programs.

Evidence for the importance of the "genotype" theory in alcohol seeking behavior: a commentary.

Consensus of the literature points towards a neuropsychogenetic model of alcoholism. Evidence in both animals and humans tends to support the proposed "genotype" theory of alcohol-seeking behavior, whereby a predisposition to alcohol preference may be mediated in part by either innate (genetic) or environmentally (stress and/or alcohol) induced brain opioid peptide dysfunction. Potential therapeutic rationale involving the utilization of novel inhibitors of carboxypeptidase A (enkephalinase) which raise endogenous enkephalin levels and possess anti-alcohol seeking effects is emphasized.

Retinal carbonic anhydrase: a comparative study.

Activity levels of carbonic anhydrase (E.C. 4.2.1.1) were measured in the retina and pigment epithelium of thirteen vertebrate species. The animals cover five taxonomic classes and among them illustrate four different retinal vascular supply patterns. The species can also be grouped according to their temperature regulatory mechanisms; i.e., homeothermic or poikilothermic. Significant differences are observed when the enzyme activities are examined by taxonomic class-birds and reptiles appear to have very high levels of enzyme activity; mammals and fish, moderate levels; and amphibia, low levels. When the enzyme activities are regrouped according to either vascular structure or temperature regulation, no significant differences are observed. From this, it is concluded that the level of carbonic anhydrase required by retinal tissue is not directly related to either of these factors. Carbonic anhydrase activity in kidney was compared with that in retina/pigment epithelium. Retina/pigment epithelium values are as great or greater than those in kidney but show a wider range. Only mammalian and avian kidney CA activity levels are high. We suggest that carbonic anhydrase levels in retina/pigment epithelium may be responsive to factors which influence vitreo-retinal ion and pH gradients.

Rapid isolation of mammalian Müller cells.

A procedure has been developed for the rapid isolation of relatively pure populations of glia, Müller cells, from mammalian retina and is reported here for the rabbit. The retina, cleaned of vitreous, is dissociated by 4 interacting operations--enzymatic digestion of extracellular matrix by means of hyaluronidase, collagenase and papain, removal of divalent cations, acidification and mild trituration. The resultant admixture consists of receptors, neurons and glia; 10% of the cells are Müller cells. These glial elements can be brought to 95% purity by rapid centrifugation on convex 0-30% Percoll gradients. Of the resultant glia, 80% exclude trypan blue. The Müller cells are enriched in two glial specific enzymes, glutamine synthetase and carbonic anhydrase and they retain a significant fraction of the membrane bound carbonic anhydrase enzyme activity. Light and scanning electron microscopy show the cells to be well preserved and covered extensively with microvilli. In the outer nuclear zone, the cells are plicated and end in bulbs tufted with microvilli. The procedure we describe allows studies of a new preparation of intact, relatively undamaged, adult, isolated mammalian Müller cells to better understand the functioning of glial cells.

Glial endocytosis of protein in the traumatized brain.

The effect of trauma on the clearance of exogenous protein from the extracellular space of the brain was examined in monkeys and rats. Manual contusion of the monkey temporal lobe or air puff contusion of the rat parietal cortex resulted in intense glial uptake of the protein marker, horseradish peroxidase (HRP). There was no evidence of punctate or diffuse material in neurons at or distant from the trauma site. Cellular uptake was evident as early as 4 hr post trauma and reached maximal expression 24 hr post injury. By 72 hr post trauma few labeled cells could be seen. The extent of the glial uptake of protein was suggested by the decrease in extracellular marker in the vicinity of the labeled cells and by calculation of the fraction of glia so labeled. Sites of labeled glia were highly localized, corresponding to regions of stress. The corpus callosum was most clearly labeled in the rat experiments. The data suggest that in cases of traumatic injury to the brain, glia may be selectively stimulated to pronounced uptake of protein marker and thus may represent a potential means of clearing the extracellular space of extravasated serum protein.

Ultrasonic decalcification of bone.

Decalcification of bone specimens of 2-5 mm thickness can be achieved in 5 hours or less when the decalcifying fluids are agitated by ultrasonic energization. The morphology of hard and soft tissue is extremely well preserved by this procedure. Acid or chelating decalcifiers may be used and the application of combined fixation-chelation permits routinely many histochemical procedures previously requiring special handling.