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Comparative study of separation of non-encapsulated drug from unilamellar liposomes by various methods.

The purpose of this study was to compare the various methods available to separate non-encapsulated drug from large unilamellar liposomes (LUV). Multilamellar liposomes (MLV) were prepared by thin film hydration using distearoylphosphatidylcholine:cholesterol (2:1 molar ratio). MLVs were passed through a 0.2 micron polycarbonate membrane using an extruder to prepare LUVs. Particle size of liposome preparations was characterized using a submicron particle-size analyser. The non-encapsulated drug was separated by: filtering through Centrifree tubes; passing through gel (Sepharose-4B and Sephadex G-25M); passing through minicolumn; ficoll density gradient; protamine aggregation; or dialysis. The dialysis method was found to be unsuitable for separation of non-encapsulated drug due to equilibration of encapsulated drug as the free drug was dialyzed. The upper limit for lipid concentration was 5 mg mL-1 using the Centrifree method. Separation using gel chromatography led to dilution of liposome preparation. Minicolumn and density gradient techniques did not lead to sample dilution, however the minicolumn method was tedious. The time required for separation of liposomes by protamine aggregation was longer for neutral liposomes. Thus it was concluded that the Centrifree was the fastest method to estimate encapsulation; the density gradient method was ideal to separate non-encapsulated drug; and protamine aggregation was the least expensive method to estimate encapsulation efficiency.

Anti-HIV Agents↗

The influence of spatial separation on divided listening.

If spatial attention acts like a "spotlight," focusing on one location and excluding others, it may be advantageous to have all targets of interest within the same spatial region. This hypothesis was explored using a task where listeners reported keywords from two simultaneous talkers. In Experiment 1, the two talkers were placed symmetrically about the frontal midline with various angular separations. While there was a small performance improvement for moderate separations, the improvement decreased for larger separations. However, the dependency of the relative talker intensities on spatial configuration accounted for these effects. Experiment 2 tested whether spatial separation improved the intelligibility of each source, an effect that could counteract any degradation in performance as sources fell outside the spatial spotlight of attention. In this experiment, intelligibility of individual sources was equalized across configurations by adding masking noise. Under these conditions, the cost of divided listening (the drop in performance when reporting both messages compared to reporting just one) was smaller when the spatial separation was small. These results suggest that spatial separation enhances the intelligibility of individual sources in a competing pair but increases the cost associated with having to process both sources simultaneously, consistent with the attentional spotlight hypothesis.

Acoustic Stimulation↗

The effect of a separate stream for minor injuries on accident and emergency department waiting times.

INTRODUCTION: To decrease waiting times within accident and emergency (A&E) departments, various initiatives have been suggested including the use of a separate stream of care for minor injuries ("fast track"). This study aimed to assess whether a separate stream of minor injuries care in a UK A&E department decreases the waiting time, without delaying the care of those with more serious injury. INTERVENTION: A doctor saw any ambulant patients with injuries not requiring an examination couch or an urgent intervention. Any patients requiring further treatment were returned to the sub-wait area until a nurse could see them in another cubicle. METHOD: Data were retrospectively extracted from the routine hospital information systems for all patients attending the A&E department for five weeks before the institution of the separate stream system and for five weeks after. RESULTS: 13 918 new patients were seen during the 10 week study period; 7117 (51.1%) in the first five week period and 6801 (49.9%) in the second five week period when a separate stream was operational. Recorded time to see a doctor ranged from 0-850 minutes. Comparison of the two five week periods demonstrated that the proportion of patients waiting less than 30 and less than 60 minutes both improved (p<0.0001). The relative risk of waiting more than one hour decreased by 32%. The improvements in waiting times were not at the expense of patients with more urgent needs. CONCLUSIONS: The introduction of a separate stream for minor injuries can produce an improvement in the number of trauma patients waiting over an hour of about 30%. If this is associated with an increase in consultant presence on the shop floor it may be possible to achieve a 50% improvement. It is recommended that departments use a separate stream for minor injuries to decrease the number of patients enduring long waits in A&E departments.

Appointments and Schedules↗

TLC separation of hippuric, mandelic, and phenylglyoxylic acids from urine after mixed exposure to toluene and styrene.

A method using thin-layer chromatography is described to determine the concentration of hippuric acid, mandelic acid, and phenylglyoxylic acid present in the urine after occupational mixed exposure to toluene and styrene. These substances are known metabolites of toluene and styrene, and therefore the evaluation to mixed exposure to toluene and styrene may be carried out separating these metabolites beforehand. Procedures are proposed to separate the metabolites as follows: (1) separation of hippuric acid from mandelic acid, (2) separation of mandelic acid from phenylglyoxylic acid, and (3) separation of hippuric acid and mandelic acid from phenylglyoxylic acid. The developing reagent p-dimethylaminobenzaldehyde in acetic acid anhydride was used after separation on Kieselgel and Silicagel. The sensitivity of the method was 6 microgram of hippuric acid, 10 microgram of mandelic acid, and 7 microgram of phenylglyoxylic acid with an average recovery of 94%.

Chromatography, Thin Layer↗

Aetiology of separation-related behaviour in domestic dogs.

A longitudinal study of seven litters of labrador retrievers and five litters of border collies from eight weeks to 18 months of age indicated that the majority showed some degree of potentially undesirable behaviour when separated from their owners. Its incidence was particularly high in the labrador retrievers, of which 13 of 23 showed separation-related behaviour for more than a month. Socially diverse environments experienced between six and 12 months of age were associated with a subsequent absence of separation-related behaviour. In a questionnaire survey of dog owners, separation-related behaviour was reported in 27 of 94 dogs, and a further 20 had shown the behaviour in the past. Male dogs were more likely to express separation-related behaviour currently, and females were more likely never to have displayed it. The prevalence of the behaviour was unaffected by whether the dog was pedigree or mixed breed, or whether it had been obtained from a breeder or from a rescue organisation. Combining the results of the two studies, the owners of only six of 75 dogs showing separation-related behaviour had sought assistance, and only two of the owners had sought help from veterinary surgeons.

Animals↗

Improved separation method for rat proximal and distal renal tubules.

We modified and improved enzyme digestion and density gradient separation procedures to obtain fractions of proximal and distal renal tubules with high yield and viability. Kidneys from two anesthetized adult Wistar rats were flushed with Krebs-Henseleit buffer (KHB) and then perfused in situ with recirculated KHB containing collagenase and hyaluronidase at 125 mmHg. Cortices were excised, minced, and incubated in KHB containing enzymes for 35 min at 37 degrees C. Dissociated tubules were removed at 10-min intervals, rinsed, and placed in KHB containing 10% calf serum, vitamins, and amino acids at 4 degrees C. Separation was achieved by suspending the tissue in 45% isosmotic Percoll layered over an undiluted Percoll cushion and centrifuging. Proximal tubules sedimented near the cushion. Distal segments were isolated in the uppermost bands of a second 35% Percoll separation. Viability was greater than 95% as measured by lactate dehydrogenase leakage and quantitated by oxygen consumption and ATP content. Basal oxygen consumption was greater than 33 nmol O2 X min-1 X mg protein-1 in all fractions and was stimulated by succinate and inhibited by amiloride and ouabain. Basal ATP content averaged 9.7 nmol/mg ATP. An average 3.3-fold separation for the proximal fraction and 24.5-fold separation for the distal fraction was assessed by the enrichment of six specific enzyme markers, with several of the markers indicating separations up to 32-fold. Isolated tubules also displayed functional responses to parathyroid hormone and vasopressin. Distal, but not proximal, segments demonstrated significantly increased adenosine 3',5'-cyclic monophosphate formation with vasopressin.

Animals↗

Spatial summation of heat-induced pain: influence of stimulus area and spatial separation of stimuli on perceived pain sensation intensity and unpleasantness.

1. Psychophysical experiments were initiated to determine the possible influence of increasing stimulus size on perceived pain intensity. Six trained human subjects (5 male, 1 female) made visual analogue scale (VAS) ratings for pain-sensation intensity and unpleasantness in response to nociceptive thermal stimuli. Test stimuli consisted of 5-s duration heat pulses (45-50 degrees C in 1 degrees increments) delivered by one, two, or three contact thermal probes (1 cm2 each) applied to the medial aspect of the anterior forearm. 2. The area of skin receiving noxious thermal stimuli was changed by randomly varying the number of thermodes activated. The effects of varying the distance between the thermal probes also were evaluated. In the first series of experiments, thermal-probe separation was kept close to 0; in subsequent experimental series, the thermodes were separated by either 5 or 10 cm. 3. In each experimental series, considerable spatial summation occurred in both pain-sensation intensity and unpleasantness dimensions of pain. This summation occurred throughout the nociceptive thermal range of 45-50 degrees C and was larger at suprathreshold temperatures (greater than or equal to 47 degrees C) than those near threshold (less than or equal to 46 degrees C). Unlike spatial summation of perceived warmth, that of pain was not characterized by systematic changes in power-function exponents but as approximately upward parallel displacements in double-logarithmic coordinates. 4. Thermal-probe separation over a range of 0-10 cm had no effects on spatial summation of pain-sensation intensity or pain unpleasantness. In contrast, increasing thermal-probe separation increased the subjects' ability to discriminate differences in stimulus size and their ability to detect correctly the number of thermal probes activated. 5. Because affective VAS ratings of unpleasantness were linearly related to, but distinctly and systematically less than, VAS ratings of pain-sensation intensity, it was clear that subjects responded quite differently to these two pain dimensions. Affective judgements were not additionally influenced by thermal probe separation and hence by the ability to perceive stimulus size or number of thermal probes activated. 6. The results indicate that powerful spatial-summation mechanisms exist for heat-induced pain. Spatial summation of pain is likely to be subserved both by local integration mechanisms at the level of single spinothalamic-tract neurons and by recruitment of central nociceptive neurons, because spatial summation of pain occurred to approximately equal extents under conditions of thermode separations over a distance of at least 20 cm.

Adult↗

Detection of auditory signals by frog inferior collicular neurons in the presence of spatially separated noise.

Detection of auditory signals by frog inferior collicular neurons in the presence of spatially separated noise. J. Neurophysiol. 80: 2848-2859, 1998. Psychophysical studies have shown that the ability to detect auditory signals embedded in noise improves when signal and noise sources are widely separated in space; this allows humans to analyze complex auditory scenes, as in the cocktail-part effect. Although these studies established that improvements in detection threshold (DT) are due to binaural hearing, few physiological studies were undertaken, and very little is known about the response of single neurons to spatially separated signal and noise sources. To address this issue we examined the responses of neurons in the frog inferior colliculus (IC) to a probe stimulus embedded in a spatially separated masker. Frogs perform auditory scene analysis because females select mates in dense choruses by means of auditory cues. Results of the extracellular single-unit recordings demonstrate that 22% of neurons (A-type) exhibited improvements in signal DTs when probe and masker sources were progressively separated in azimuth. In contrast, 24% of neurons (V-type) showed the opposite pattern, namely, signal DTs were lowest when probe and masker were colocalized (in many instances lower than the DT to probe alone) and increased when the two sound sources were separated. The remaining neurons demonstrated a mix of these two types of patterns. An intriguing finding was the strong correlation between A-type masking release patterns and phasic neurons and a weaker correlation between V-type patterns and tonic neurons. Although not decisive, these results suggest that phasic units may play a role in release from masking observed psychophysically. Analysis of the data also revealed a strong and nonlinear interaction among probe, masker, and masker azimuth and that signal DTs were influenced by two factors: 1) the unit's sensitivity to probe in the presence of masker and 2) the criterion level for estimating DT. For some units, it was possible to examine the interaction between these two factors and gain insights into the variation of DTs with masker azimuth. The implications of these findings are discussed in relation to signal detection in the auditory system.

Acoustic Stimulation↗

Comparison of different protocols in plateletpheresis with the haemonetics MCS 3p blood cell separator with respect to parameters of product quality.

BACKGROUND: In thrombocytapheresis, intermittent flow cell separators produce platelet concentrates (PC) with a higher leucocyte contamination than continuous flow blood cell separators. The new discontinuous flow cell separator Haemonetics MCS 3p offers a low-leucocyte solution for PC. The quality of PC obtained by the MCS 3p was investigated in this study. DESIGN: Prospective study. SETTING: Haemapheresis Unit of a University Clinic. PATIENTS: Healthy blood donors from the haemapheresis unit. MATERIALS AND METHODS: Platelet (PLT) yield, separation efficiency and white blood cell (WBC) contamination were studied in three different protocols. Two protocols used a haemocalculator, which calculated the target volume based on the donor's physical characteristics and the desired PLT yield for the procedure. Protocol I used 3,000 ml as target process volume and protocol II 3.3 x 10(11) as desired PLT yield. Protocol III was used without haemocalculator. Glucose, lactate, lactate dehydrogenase (LDH), morphology score and pH value were analysed to investigate the quality of the PC. RESULTS: Platelet yield and separation efficiency were not statistically different in the three protocols. Leucocyte contamination was lowest in the protocol without haemocalculator (median: 3.15 x 10(6), range 0.4-20.8 x 10(6)). Glucose, lactate, LDH and pH were not statistically different in the three protocols. Morphology score was best in protocol III. CONCLUSIONS: PLT collection with the MCS 3p blood cell separator results in sufficient thrombocyte yields. Using the haemocalculator we were not able to achieve the desired platelet yield. For this reason, and because of the higher WBC contamination in protocol II we prefer PLT collection without the haemocalculator. The quality of the platelet concentrates was good with respect to the parameters glucose, lactate, LDH, morphology score and pH.

Adult↗

Flow separation in the renal arteries.

To determine the nature of the flow in the proximal portion of the renal arteries, a common site for the development of atherosclerosis, we performed pulsatile flow studies in a clear acrylic mold of a normal human aorta and renal arteries. The mold was developed from a cast of the renal arteries prepared in situ during the autopsy of a 27-year-old woman. Flow in the mold was visualized by the illumination of bouyant particles (100 to 300 mu). A range of branch (renal) to trunk (aorta) flow ratios of 0.053 to 0.350 was studied. Flow separation during systole was considered present when particles near the wall reversed direction. Flow separation occurred throughout systole at the superior aspect of the origin of both the left and right renal arteries at a branch-to-trunk flow ratio of 0.053. As the branch-to-trunk flow ratio increased, flow separation occurred during the deceleration phase of right renal flow. At a branch-to-trunk flow ratio of 0.350, flow separation was no longer present. This suggests that flow separation may occur near the renal ostia if renal flow falls appreciably while aortic flow remains elevated. Because flow separation involves low wall shear, mass transfer across the arterial wall may be adversely affected and possibly contribute to the formation of atheroma.

Adult↗

Resonance Raman spectroscopy characterization of single-wall carbon nanotube separation by their metallicity and diameter.

Several techniques were recently reported for the bulk separation of metallic (M) and semiconducting (S) single wall carbon nanotubes (SWNTs), using optical absorption and resonance Raman spectroscopy (RRS) as a proof of the separation. In the present work, we develop a method for the quantitative evaluation of the M to S separation ratio, and also for the SWNT diameter selectivity of the separation process, based on RRS. The relative changes in the integrated intensities of the radial-breathing mode (RBM) features, with respect to the starting material, yield the diameter probability distribution functions for M and S SWNTs in the separated fractions, accounting for the different resonance conditions of individual SWNTs, while the diameter distribution of the starting material is obtained following the fitting procedure developed by Kuzmany and coworkers. Features other than the RBM are generally less effective for characterization of the separation process for SWNTs.

Metals↗

Thresholds for stereo-slant discrimination between spatially separated targets are influenced mainly by visual and memory factors but not oculomotor instability.

Surface-slant variations can be sensed either simultaneously with steady fixation or sequentially with saccadic gaze shifts. Stereo-slant discrimination thresholds are affected by visual, oculomotor, and memory factors. We have investigated the effects of fixation strategy, target separation, and exposure duration on stereo-slant discrimination. With long exposure durations (734 ms), stereo-slant discrimination thresholds measured with simultaneous presentation of test and reference stimuli were lower with gaze shifts than without them when target separations exceeded 4 deg. Above 4-deg target separations, the benefits of improved disparity resolution with foveal gaze shifts outweighed the costs of oculomotor variability associated with saccades. With short exposure durations (167 ms), as target separation increased, stereo-slant discrimination thresholds measured without gaze shifts increased with both sequential and simultaneous stimulus presentations, whereas thresholds with gaze shifts remained constant. This indicates that oculomotor errors are not an important factor in stereo-slant discrimination. In contrast to stereo-slant thresholds, sequential stereo-depth thresholds between two dots, measured with gaze shifts, increased with target separation. Thus, oculomotor error increases with target separation, and it is an important factor in stereo-depth discrimination.

Depth Perception↗

The occupational burden of mental disorders in theU.S. military: psychiatric hospitalizations, involuntary separations, and disability.

OBJECTIVE: A recent study showed that among U.S. military personnel, mental disorders were the leading medical correlate of separation from military service. The reasons for this association have not been determined. The purpose of this study was to characterize the risk and reasons for service separation for soldiers hospitalized with mental disorders compared with those hospitalized for other illnesses. METHOD: Population-based electronic health care data were linked with data on separations and disability. The authors constructed a cohort of 13,971 U.S. Army soldiers first hospitalized in 1998 and followed them for up to 2 years following this first hospitalization. RESULTS: The rate of service separation 6 months after first hospitalization was 45% among personnel whose primary hospital discharge diagnosis was a mental disorder, 27% among those with a secondary mental disorder discharge diagnosis, and 11% among those hospitalized for all other medical conditions. Separation from military service due to medical disability, misconduct and other legal problems, unauthorized work absences, behavior related to personality disorders, and alcohol rehabilitation failure was significantly more common following hospitalization for a mental disorder than for other medical conditions. Mental disorders were also significantly associated with medical separations in which there was evidence that the condition existed prior to service. CONCLUSIONS: In the military, the occupational impact of mental disorders compared with other medical conditions appears to be mediated not only by greater disease chronicity and severity but also by a variety of behavioral problems including misconduct, legal problems, unauthorized absences, and alcohol/drug-related problems. The study also points to the difficulties inherent in screening for mental disorders prior to entry into military service.

Cohort Studies↗

Separation-individuation theory and attachment theory.

Separation-individuation and attachment theories are compared and assessed in the context of psychoanalytic developmental theory and their application to clinical work. As introduced by Margaret Mahler and John Bowlby, respectively, both theories were initially regarded as diverging from traditional views. Separation-individuation theory, though it has had to be corrected in important respects, and attachment theory, despite certain limitations, have nonetheless enriched psychoanalytic thought. Without attachment an infant would die, and with severely insecure attachment is at greater risk for serious disorders. Development depends on continued attachment to a responsive and responsible caregiver. Continued attachment to the primary object was regarded by Mahler as as intrinsic to the process of separation-individuation. Attachment theory does not account for the essential development of separateness, and separation-individuation is important for the promotion of autonomy, independence, and identity. Salient historical and theoretical issues are addressed, including the renewed interest in attachment theory and the related decline of interest in separation-individuation theory.

Adolescent↗

Effect of litter separation on 24-hour rhythmicity of plasma prolactin, follicle-stimulating hormone and luteinizing hormone levels in lactating rabbit does.

BACKGROUND: This work describes the effect of a 48-h litter separation on 24-h patterns of plasma prolactin, FSH and LH concentration in female lactating rabbits kept under a 16:8 light-dark photoperiod (lights on at 0800 h). METHODS: Groups of 6-7 female lactating rabbits maintained with their litters or separated from them for 48 h were killed by decapitation on day 11 post-partum, at 6 different time points throughout a 24-h cycle, starting at 0900 h. Plasma levels of prolactin, FSH and LH were measured by specific double antibody radio-immunoassays. RESULTS: Plasma level of prolactin in control and separated does changed in a similar way throughout the day, showing two maxima, at 0500-0900 h and at 1700-2100 h, respectively. Litter separation significantly augmented plasma FSH and LH and disrupted their 24-h rhythmicity. CONCLUSION: Since previous studies had shown that litter separation for short periods of time augmented sexual receptivity and fertility of the doe, the changes in FSH and LH reported may influence the massive release of gonadotropin releasing hormone, LH and FSH triggered by mating or artificial insemination in litter-separated mothers.

Journal Article↗

Childhood separation experiences and psycho-social status in primiparous women: preliminary findings.

In a group of British-born primiparous women, admission of a separation experience from parents in childhood was found to be associated with unmarried status, teenage pregnancy, having had psychiatric treatment, housing problems and a high score on a 'Malaise' inventory. Within the separated group the unmarried and teenage status were associated only with separations occurring in the context of a disrupted childhood, but were not related to the age of the separations. The Malaise score, however, was raised both for disrupted separations and for those occurring below the age of 5. It is noted that the factors associated with the separation in the mothers are those which have been found to be more common in the mothers of children with behaviour disorders.

Adolescent↗

Incomplete sister chromatid separation is the mechanism of programmed chromosome elimination during early Sciara coprophila embryogenesis.

Sex in Sciara coprophila is determined by maternally supplied factors that control the number of paternal X chromosomes eliminated during the syncytial embryonic divisions. Confocal microscopy and FISH demonstrate that the centromeres of the X chromosomes separate at anaphase and remain functional during the cycle in which the X chromosomes are eliminated. However, a region of the sister chromatids fails to separate and the X chromosomes remain at the metaphase plate. This indicates that failure of sister chromatid separation is the mechanism of chromosome elimination. Elimination of the X chromosomes requires the presence of a previously discovered Controlling Element that acts in cis during male meiosis. Using an X-autosome translocation, we demonstrate that the Controlling Element acts at-a-distance to prevent sister chromatid separation in the arm of an autosome. This indicates that the region in which sister chromatid separation fails is chromosome-independent. Although chromosome elimination occurs in all somatic nuclei and is independent of location of the nuclei within the embryo, the decision to eliminate is made at the level of the individual nucleus. Programmed X chromosome elimination occurs at different cycles in male and female embryos. These observations support a model in which elements on the X chromosome are titrating maternally supplied factors controlling the separation of sister X chromatids.

Animals↗

The endo-beta-1,3-glucanase eng1p is required for dissolution of the primary septum during cell separation in Schizosaccharomyces pombe.

Schizosaccharomyces pombe cells divide by medial fission throughout contraction of an actomyosin ring and deposition of a multilayered division septum that must be cleaved to release the two daughter cells. Although many studies have focused on the actomoysin ring and septum assembly, little information is available concerning the mechanism of cell separation. Here we describe the characterization of eng1+, a new gene that encodes a protein with detectable endo-beta-1,3-glucanase activity and whose deletion is not lethal to the cells but does interfere in their separation. Electron microscopic observation of mutant cells indicated that this defect is mainly due to the failure of the cells to degrade the primary septum, a structure rich in beta-1,3-glucans, that separates the two sisters cells. Expression of eng1+ varies during the cell cycle, maximum expression being observed before septation, and the protein localizes to a ring-like structure that surrounds the septum region during cell separation. This suggests that it could also be involved in the cleavage of the cylinder of the cell wall that covers the division septum. The expression of eng1+ during vegetative growth is regulated by a C2H2 zinc-finger protein (encoded by the SPAC6G10.12c ORF), which shows significant sequence similarity to the Saccharomyces cerevisiae ScAce2p, especially in the zinc-finger region. Mutants lacking this transcriptional regulator (which we have named ace2+) show a severe cell separation defect, hyphal growth being observed. Thus, ace2p may regulate the expression of the eng1+ gene together with that of other genes whose products are also involved in cell separation.

Base Sequence↗