Molecular dynamics study of surface-tethered S(CH2CH2O)6CH3: helix formation and thermal disorder.

We present the results of a molecular dynamics study of a set of surface-tethered S(CH2CH2O)6CH3 chains. In this study, we analyze helix formation, in addition to thermal disorder, and find that spontaneous helix formation and details of helix morphology depend on charge partitioning ascribed to oxygen and the methylene groups. The effects of varying surface coverage as well as chain-surface interaction strength indicate that a set of approximately 7/2 helical structures oriented predominantly normal to the surface are formed at near full coverage. This occurs even though thermal disorder clearly precludes a description based on the concept of a perfect crystalline monolayer. Thermal fluctuations in chain morphology in the vicinity of the terminal methyl groups lead to the exposure of oxygen to the external environment. We also find that the persistence of compact helix-containing domains at partial surface coverage results in the formation of well-defined cavities or void regions that expose the bare surface, even in the presence of strong chain-surface attractive interactions.

Increasing incidence of admissions to a General Hospital for deliberate self-harm in trinidad / Aumento de la incidencia de ingresos al Hospital General por auto-daño deliberado en trinidad

BACKGROUND: Deliberate self-harm and suicidal behaviour have become an increasingly common form of morbidity in the developing world. Suicidal behaviour is a major public health problem in Trinidad as the country has the second highest completed suicide rate in the English-speaking Caribbean. The objectives of this study were to determine the epidemiology of attempted suicide and self-harm at a specified site (the Port-of-Spain General Hospital) and compare it to previous studies done at the same site. SUBJECTS AND METHODS: This was investigated through a review of one years' admissions to the Port-of-Spain General Hospital for suicidal behaviour. Incidence was compared with a previous study completed at this site and reported in 1974. Comparison of the demographic characteristics of the sample with that of the previous study was also undertaken using chi-square analysis and significance testing through the use of t tests. RESULTS: A total of 368 referrals were made for attempted suicide or deliberate self-harm over the period indicating a fourfold increase in the incidence of this behaviour with a greater increase among males where the female to male ratio has declined from 4 to 1 to 2 to 1, p < 0.001; the mean age of males was 34 years compared to females 22 (p < 0.0001). The males were more likely to use violent means compared to females who were more likely to ingest tablets or bleach. Relationship difficulties were most commonly cited by both males and females as the reason for their attempt. There was a trend to greater propensity for this behaviour among Indo-Trinidadians as compared to Afro-Trinidadians in both females and males. CONCLUSIONS: Increasing numbers of men are engaging in self-harm behaviour and are using more violent and physically harmful methods suggesting a greater degree of suicidal risk while women mainly engage in acts of ingestion with a much lower risk of death. The older mean age of these men suggests that their problems are presenting in middle adulthood while women are engaging in this behaviour in young adulthood. Suicidal behaviour or deliberate self-harm is a major public health problem in Trinidad.

ANTECEDENTES: El auto-daño deliberado y el comportamiento suicida se han convertido cada vez más en una forma común de morbosidad en el mundo en vías de desarrollo. El comportamiento suicida es un serio problema de salud pública en Trinidad, como país que tiene la tasa más alta de suicidios efectuados en el Caribe anglófono. Los objetivos de este estudio fueron determinar la epidemiología del suicidio efectuado y el auto-daño en un lugar específico (el Hospital General de Puerto España) y compararla con estudios previos realizados en el mismo lugar. SUJETOS Y MÉTODOS: Esto fue investigado mediante una revisión de los ingresados de un año al Hospital General de Puerto España por comportamiento suicida. La incidencia fue comparada con un estudio previo terminado en este sitio y reportado en 1974. La comparación de las características demográficas de la muestra con la del estudio previo se llevó a cabo usando un análisis de chi-cuadrado y pruebas de significatividad mediante el uso de tests t. RESULTADOS: Se produjeron un total de 368 casos referidos por intento de suicidio o auto-daño deliberado en el período en cuestión, lo cual indica que la incidencia de esta conducta aumentó en cuatro veces, con un mayor incremento entre los varones, y una disminución de la proporción hembras-varones de 4:1 a 2:1; p < 0.001); la edad promedio de los varones fue 34 años en comparación con la de las hembras, 22 (p < 0.0001). Los varones mostraron una tendencia mayor a usar medios violentos en comparación con las hembras, entre las que había una mayor probabilidad de ingerir tabletas o lejía. Las dificultades en las relaciones constituyeron la explicación más comúnmente dada como causa del intento, tanto por los varones como por las hembras. Hubo tendencia a una mayor propensión a este comportamiento entre los indo-trinitenses en comparación con los afro-trinitenses, tanto en hembras como varones. CONCLUSIONES: Un número cada vez mayor de hombres se involucran en conductas auto-agresivas y están usando métodos más violentos y físicamente dañinos, lo que sugiere un grado mayor de riesgo de suicidio, mientras que las mujeres se entregan principalmente a actos de ingestión con riesgo mucho menor de muerte. La edad promedio mayor de estos hombres sugiere que sus problemas se presentan en medio de su adultez, mientras que las mujeres se entregan a estas conductas como adultos jóvenes. La conducta suicida y el auto-daño deliberado constituyen un problema principal de salud pública en Trinidad.

Can machine perfusion decrease the likelihood of discard among biopsied kidneys?

Many factors, such as donor risk factors and renal function, have been shown to be associated with an increased likelihood of discard after recovering kidneys from deceased donors. When these factors are insufficient for assessment, renal biopsy is often performed at the time of harvest to assess suitability. Our aims were to identify factors that predict the discard of a biopsied kidney and to assess the impact of machine perfusion (MP) on kidney discard. We biopsied 678 kidneys from deceased donors aged >or=40 years from 2001 to 2006. We used a logistic regression model to estimate the adjusted odds ratios for kidney discard. Thirty-nine percent (n = 261) of biopsied kidneys were discarded. Kidneys with glomerulosclerosis (GS) > 20% had the highest likelihood of discard. Other significant predictors of discard included extreme donor age, final resistance (>40), atherosclerosis, interstitial fibrosis, arteriolosclerosis, and terminal serum creatinine value (SCr) > 1.5 mg/dL. MP kidneys (n = 69) were less likely to be discarded than cold storage (CS) kidneys after adjusting for other factors (adjusted odds ratio = .13, P < .001). In conclusion, abnormal biopsy findings were associated with the highest likelihood of discard. MP was used in only 10% of the cases; however, the use of MP was associated with a decreased likelihood of discard among biopsied kidneys.

Factors associated with discard of recovered kidneys.

We reviewed diseased donor (DD) kidney usage at a single Organ Procurement Organization in Southern California to more closely examine factors associated with discard. From 2001 to 2006, 3863 kidneys from 1959 DDs were recovered, but 454 (11.8%) were subsequently discarded. Among the discarded kidneys, 211 (46.5%) were discarded based upon biopsy findings, 19 (4.2%) due to anatomical abnormalities, 16 (3.5%) based on donor quality, and 14 (3.1%) because they were felt to be too old to be pumped. Multivariate logistic regression analysis was performed using significant prognostic factors upon univariate analyses. According to the magnitude of the adjusted odds ratio (AOR), significant prognostic factors for discard were extreme donor age (AOR = 24.1 of age 70-80 years, P < .001; AOR = 6.34 age 50-69 years, P < .001; AOR = 2.77 age 40-49 years, P < .001; and AOR = 3.09 age <10 years, P < .001 vs age 10-39 years), high final resistance (AOR = 8.86 of >40 vs others, P = .006), glomerulosclerosis (GS) > 20% (AOR = 5.94 vs GS 0%-5%/no biopsy, P < .001), severe atherosclerosis (AOR = 4.66, P = .003), abnormal anatomy (AOR = 2.7, P < .001), and moderate or severe arteriolosclerosis (AOR = 2.2 vs none/mild/no biopsy, P < .001). Among biopsy findings, the presence of GS > 20% was associated with the highest likelihood of discard. A high final resistance increased the likelihood of discard as well. In conclusion, these findings may help to set the groundwork toward a more uniform approach to organ utilization in donor service areas.

Increasing incidence of admissions to a general hospital for deliberate self-harm in Trinidad.

BACKGROUND: Deliberate self-harm and suicidal behaviour have become an increasingly common form of morbidity in the developing world Suicidal behaviour is a major public health problem in Trinidad as the country has the second highest completed suicide rate in the English-speaking Caribbean. The objectives of this study were to determine the epidemiology of attempted suicide and self-harm at a specified site (the Port-of-Spain General Hospital) and compare it to previous studies done at the same site. SUBJECTS AND METHODS: This was investigated through a review of one years' admissions to the Port-of-Spain General Hospital for suicidal behaviour. Incidence was compared with a previous study completed at this site and reported in 1974. Comparison of the demographic characteristics of the sample with that of the previous study was also undertaken using chi-square analysis and significance testing through the use of t tests. RESULTS: A total of 368 referrals were made for attempted suicide or deliberate self-harm over the period indicating a fourfold increase in the incidence of this behaviour with a greater increase among males where the female to male ratio has declined from 4 to 1 to 2 to 1, p < 0.001; the mean age of males was 34 years compared to females 22 (p < 0.0001). The males were more likely to use violent means compared to females who were more likely to ingest tablets or bleach. Relationship difficulties were most commonly cited by both males and females as the reason for their attempt. There was a trend to greater propensity for this behaviour among Indo-Trinidadians as compared to Afro-Trinidadians in both females and males. CONCLUSIONS: Increasing numbers of men are engaging in self-harm behaviour and are using more violent and physically harmful methods suggesting a greater degree of suicidal risk while women mainly engage in acts of ingestion with a much lower risk of death. The older mean age of these men suggests that their problems are presenting in middle adulthood while women are engaging in this behaviour in young adulthood. Suicidal behaviour or deliberate self-harm is a major public health problem in Trinidad.

Awareness of potential valvulopathy risk with pergolide and changes in clinical practice after label change: a survey among European neurologists.

In response to concern about the reported frequency of ergot-associated valvulopathy in patients with Parkinson's disease (PD), Eli Lilly and Company updated the Risk Minimization Program for pergolide, changing the Summary of Product Characteristics (SmPC) and distributing a Dear Doctor Letter (DDL) highlighting the new label changes. A survey was conducted subsequently to assess neurologists' awareness of the revised SmPC and their resulting changes in practice. A random sample of 20.3% of neurologists (n = 4056) from 12 eligible EU countries were invited to participate. Of the target population of 247 neurologists who treated patients with PD, used pergolide in 2005, and were willing to participate, 244 (99%) responded. Overall awareness of the DDL and the SmPC changes was 94.2%. Over half (58.3%) of neurologists indicated that they prescribe pergolide exclusively as second-line treatment, although some (21.9%) used pergolide exclusively as first-line treatment. In response to the DDL, most neurologists perform echocardiograms before treatment (67.5%) and during treatment (76.7%), and over half (55%) avoid prescribing doses >5 mg/day. Overall, use of a DDL to communicate an SmPC change was effective in increasing the awareness of pergolide-associated valvulopathy and in modifying neurologists' clinical practice to minimize this risk.

Preliminary bioassay of Neem (Azadirachta indica) bark extract as a phytoacaricide against test species Boophilus microplus

This paper describes the extraction of the Neem (Azadirachta indica) bark and the determination of any statistically significant ascaricidal activity in the extract. The hexane extract of a bark sample, after reconstitution in dimethlysufoxide, was used to perform a bioassay on tick (Boophilus microplus) specimens, which were collected from cattle that had not been exposed to any acaricide for six weeks preceding (tick) collection. The percentage mortality of Boophhilus microplus obtained post administration were 100% for the pyrethroid cypermethrin, 56% for Azadirachta indica bark extract, distilled water- 8% and dimethylsulfoxide-16%. These differences in results were statistically significant at p<0.001.

Preliminary bioassay of Neem (Azadirachta indica) bark extract as a phytoacaricide against test species Boophilus microplus

This paper describes the extraction of the Neem (Azadirachta indica) bark and the determination of any statistically significant ascaricidal activity in the extract. The hexane extract of a bark sample, after reconstitution in dimethlysufoxide, was used to perform a bioassay on tick (Boophilus microplus) specimens, which were collected from cattle that had not been exposed to any acaricide for six weeks preceding (tick) collection. The percentage mortality of Boophhilus microplus obtained post administration were 100% for the pyrethroid cypermethrin, 56% for Azadirachta indica bark extract, distilled water- 8% and dimethylsulfoxide-16%. These differences in results were statistically significant at p<0.001.

Molecular dynamics study of the surface tension of a binary immiscible fluid.

The planar interface between two liquids having two degrees of affinity to mix has been studied by molecular dynamics simulations. The surface tension is calculated from the normal, PN, and transverse, PT, components of the pressure tensor P for a wide range of temperatures. An unusual increase in surface tension with increasing temperature is attributed to a pressure induced void transfer mechanism that is justified by basic thermodynamic arguments. This effect is diminished on the addition of a modest attractive potential between the two species, and there is a turnover point at higher temperatures beyond which the surface tension decreases with increasing temperature. An order parameter is identified as the gradient of the mole fraction distribution through the interfacial region. An additional effect is the dramatic inversion of the kinetic and potential contributions to the PN profile as the temperature is varied. It is found that a commonly used approximation for P, the Irving-Kirkwood 1 or IK1 method, results in a relatively modest unphysical variability in PN that weakly violates the condition of local mechanical stability. However, this artifact does not prevent the IK1 method from producing an interfacial tension which is nearly identical to that derived from the complete IK formula with no additional approximations.

Patterns of use of sildenafil among commercially insured adults in the United States: 1998-2002.

Sildenafil is increasingly being marketed to younger healthcare consumers. The purpose of this study was to profile sildenafil use in commercially insured, adult beneficiaries. Annual ambulatory prescription claims data from 1998 to 2002, for a nationwide, random sample of over 5 million life-years of commercially insured adults (aged > or =18 y), were examined retrospectively. The overall prevalence of sildenafil use increased from 0.8% (1998) to 1.4% (2002), an 84% increase. While the growth in use slowed in older males, use became more pronounced in younger males and females and decreased in older females. The fastest growing segment of users was found to be males aged 18-45 y. The proportion of users who had two or more claims for a medication that is suspected of inducing erectile dysfunction (ED) and/or a marker for a suspected ED-inducing disease decreased over the study period. Our findings suggest that use may increase among younger male and female patients and those without an underlying etiologic reason for use.

Preliminary determination of median lethal dose of Azadirachta indica (A. ) Juss bark extract against Boophilus microplus

The study was carried out in order to determine the effective dose of Azadirachta indica as an acaride. From a range of effective doses, nine (9) test doses ranging from zero (0) parts per million to one hundred and fifty parts (150) per million were selected. The test species Boophilus microplus was subjected to a standard bioassay. A dose response curve was plotted of percentage mortality of B. microplus against logarithmic dosage of A indica. The lethal dose, LD50 and LD100 were also determined. The LD50 was 30.20 parts per million and LD100 was found to lie between 80 and 100 parts per million. Regression analysis was performed to verify correlation of data. The data proved to be linear with a correlation coefficient of 0.89. The results of this study indicate that when the dose required to achieve 50 percent mortality is compared to average yield of A. indica bark extract obtained in previous studies (Maharaj et al 2003), the extract appears to be feasible with respect to production for use as a prospective pesticide. This is an important point for consideration as the failure in the commercialization of many plant products is often due to the yields being too low. The bark extract of Azadirachta indica has thus far proven to be worthy of consideration for further development as a phytoacaricide

A retrospective study of the treatment of toad poisoning in dogs in Trinidad.

A retrospective study of 143 cases of dogs poisoned by toads in Trinidad revealed that for the 81 cases for which the clinical signs were recorded, cardiac toxicity may have been missed due to the aggressive use of atropine therapy and/or lack of equipment. The most common clinical signs were hypersalivation and vomiting (31 reports) and convulsive seizures (14) - each ascribed to the effects of certain components present in the toad's glandular secretions (producing irritation and neurotoxicity); and hyperthermia (11). For 42 cases, treatment records indicated that atropine was the favoured pharmacological intervention (30 instances), followed by corticosteroids (20), antibiotics (19), benzodiazepines (15) and fluid therapy (14). These treatment strategies, based on symptomatic support, seemed to be successful even though they appeared to lack coherence. Of the 90 cases for which outcome data were available, 85 dogs recovered fully and 5 died. Early attention to cardiac abnormalities, using electrocardiograms, is required especially in severe cases. The use of propranolol is also warranted in these cases. The standard management of toad poisoning must also include decontamination of the gastrointestinal tract involving the washing of the mouth and the use of adsorbents. The usefulness of corticosteroids and antibiotic therapies should be re-examined.

A retrospective study of the treatment of toad poisoning in dogs in Trinidad.

A retrospective study of 143 cases of dogs poisoned by toads in Trinidad revealed that for the 81 cases for which the clinical signs were recorded, cardiac toxicity may have been missed due to the aggressive use of atropine therapy and/or lack of equipment. The most common clinical signs were hypersalivation and vomiting (31 reports) and convulsive seizures (14) - each ascribed to the effects of certain components present in the toad's glandular secretions (producing irritation and neurotoxicity); and hyperthermia (11). For 42 cases, treatment records indicated that atropine was the favoured pharmacological intervention (30 instances), followed by corticosteroids (20), antibiotics (19), benzodiazepines (15) and fluid therapy (14). These treatment strategies, based on symptomatic support, seemed to be successful even though they appeared to lack coherence. Of the 90 cases for which outcome data were available, 85 dogs recovered fully and 5 died. Early attention to cardiac abnormalities, using electrocardiograms, is required especially in severe cases. The use of propranolol is also warranted in these cases. The standard management of toad poisoning must also include decontamination of the gastrointestinal tract involving the washing of the mouth and the use of adsorbents. The usefulness of corticosteroids and antibiotic therapies should be re-examined.

Site-directed mutation of conserved cysteine residues does not inactivate the Streptococcus pyogenes hyaluronan synthase.

Hyaluronan synthase (HAS), the enzyme responsible for the production of hyaluronic acid (HA), is a well-conserved membrane-bound protein in both prokaryotes and eukaryotes. This enzyme performs at least six discrete functions in producing a heterodisaccharide polymer of several million molecular weight and extruding it from the cell. Among the conserved motifs and domains within the Class I HAS family are four cysteine residues. Cysteines in many proteins are important in establishing and maintaining tertiary structure or in the coordination of catalytic functions. In the present study we utilized a combination of site-directed mutagenesis, chemical labeling, and kinetic analyses to determine the importance of specific Cys residues for catalysis and structure of the HA synthase from Streptococcus pyogenes (spHAS). The enzyme activity of spHAS was partially inhibited by cysteine-reactive chemical reagents such as N-ethylmaleimide. Quantitation of the number of Cys residues modified by these reagents, using MALDI-TOF mass spectrometry, demonstrated that there are no stable disulfide bonds in spHAS. The six Cys residues of spHAS were then mutated, individually and in various combinations, to serine or alanine. The single Cys-mutants were all kinetically similar to the wild-type enzyme in terms of their V(max) and K(m) values for HA synthesis. The Cys-null mutant, in which all Cys residues were mutated to alanine, retained approximately 66% of wild-type activity, demonstrating that despite their high degree of conservation within the HAS family, Cys residues are not absolutely necessary for HA biosynthesis by the spHAS enzyme.

Selection and implementation for coagulation instruments/reagents in a multiple hospital/clinic network.

Selection, standardization, and implementation of instrumentation and reagents throughout a health care facility network can often be a difficult process. However, in today's ever-changing health care setting, it is often mandated. The Veteran's Integrated Systems Network 16 (VISN 16) was faced with such a task early in 1999, with the targeted area being its coagulation laboratories. The plan outlined in this paper was drafted to help facilitate the selection, standardization and implementation of coagulation systems for 17 health care facilities that make up the VISN 16 network. The VISN, encompassing 170,000 square miles, has 10 tertiary care hospitals, six of which have close relationships with affiliate universities. There are 299,733 patients enrolled in this health delivery system. The facilities range from large institutions performing both tertiary and outpatient care to small outpatient clinics. Because of the plan's detailed, comprehensive content, which included analyses of a large number of performance parameters as well as cost-efficiency, the selection process was carried out using a checklist that could be helpful to other organizations selecting equipment and reagents for coagulation studies. An implementation process was devised, resulting in coagulation standardization across the Integrated Health Network.

Identifying agent(s) associated with poult enteritis mortality syndrome: importance of the thymus.

Poult enteritis mortality syndrome (PEMS), a highly infectious disease of young turkeys, causes serious financial losses to the turkey industry. Clinically, PEMS is defined by mortality profiles, diarrhea, growth depression, and immunosuppression. Although many viruses, bacteria, and parasites are found in PEMS-infected birds, the inciting agent remains unknown. Experimentally, PEMS can be reproduced by exposing naïve poults to the intestinal contents from infected birds. Previous reports suggest that extraintestinal tissues fail to reproduce the disease. Histopathologic examination of tissues from PEMS-infected poults suggested that the thymus exhibited the earliest signs of pathology. On the basis of these observations, we hypothesized that the thymus harbors an agent(s) involved in PEMS. In these studies, naïve turkey poults were orally inoculated with a bacteria-free filtrate composed of either the intestines and feces or the thymus from PEMS-infected birds and were monitored for clinical signs of PEMS. Poults exposed to a filtrate composed solely of the thymus from PEMS-infected birds exhibited diarrhea, growth depression, mortality, pathology, and, most importantly, immunosuppression similar to poults exposed to the intestinal filtrate. The results of this study suggest that the thymus of infected birds harbors the agent(s) that can reproduce a PEMS-like disease in turkey poults.

Homicide by paraquat poisoning.

We report on two cases of homicide by paraquat poisoning. Both cases occurred in children and the common clinical presentations were gastrointestinal ulceration and acute respiratory distress, with pneumomediastinitis. Bullous emphysema was a common autopsy finding. Physicians and pathologists should be aware of these findings, for they are almost diagnostic of acute paraquat poisoning.

Forensic analysis of acute fatal poisonings in the southern districts of Trinidad.

A prospective autopsy study was undertaken at General Hospital in San Fernando, Trinidad, to analyze deaths that occurred from poisoning during 1996 and 1997. During that period 105 deaths occurred from acute poisoning. The major poisons used were: paraquat (80 cases), organophosphate/carbamate insecticides (10 cases), and anti-psychotic drugs (6 cases). Much less frequently, battery acid, ETHREL[Ethephon; (2-chloroethyl)phosphonic acid], phenols, ethanol, kerosene and flavine were the agents of choice. Suicide accounted for 99 (94.29%) deaths. Of these, 44.44% occurred in the 10-29 y-age-group. Ingestion of paraquat seems almost always fatal, since the large volumes ingested make treatment universally ineffective. Aspiration pneumonitis (100% of cases), and pneumothorax with pneumomediastinum (18.75% of cases), were remarkable autopsy findings in those dying from paraquat poisoning, while asphyxia due to respiratory paralysis was the mode of death from pesticide ingestion. Efforts to prevent suicide by poisoning may be more useful than treatment protocols.

Purification and lipid dependence of the recombinant hyaluronan synthases from Streptococcus pyogenes and Streptococcus equisimilis.

The two hyaluronan synthases (HASs) from Streptococcus pyogenes (spHAS) and Streptococcus equisimilis (seHAS) were expressed in Escherichia coli as recombinant proteins containing His6 tails. Both enzymes were expressed as major membrane proteins, accounting for approximately 5-8% of the total membrane protein. Using nickel chelate affinity chromatography, the HASs were purified to homogeneity from n-dodecyl beta-D-maltoside extracts. High levels of HAS activity could be achieved only if the purified enzymes were supplemented with either bovine or E. coli cardiolipin (CL), although bovine CL gave consistently greater activity. Mass spectroscopic analysis revealed that the fatty acid compositions of these two CL preparations did not overlap. The two HAS enzymes showed similar but distinct activation profiles with the 10 other lipids tested. For example, phosphatidic acid and phosphatidylethanolamine stimulated seHAS, but not spHAS. Phosphatidylserine stimulated both enzymes. spHAS appears to be more CL-specific than seHAS, although both purified enzymes still contain endogenous CL that can not easily be removed. Both seHAS and spHAS were inhibited by phosphatidylcholine, sphingomyelin, and sulfatides and were not substantially stimulated by cerebrosides, phosphatidylglycerol, or phosphatidylinositol. With both HASs, CL increased the Km for UDP-GlcUA, but decreased the Km for UDP-GlcNAc and gave an overall stimulation of Vmax. A kinetic characterization of the two membrane-bound and purified HASs is presented in the accompanying paper (Tlapak-Simmons, V. L., Baggenstoss, B. A., Kumari, K., Heldermon, C., and Weigel, P. H. (1999) J. Biol. Chem. 274, 4246-4253). Both purified HASs became inactive after storage for approximately 5 days at 4 degreesC. Both purified enzymes also lost activity over 4-5 days when stored at -80 degreesC in the presence of CL, but reached a level of activity that then slowly decreased over a period of months. Although the purified enzymes stored in the absence of CL at -80 degreesC were much less active, the enzymes retained this same low level of activity for at least 5 weeks. When both spHAS and seHAS were stored without CL at -80 degreesC, even after 2 months, they could be stimulated by the addition of bovine CL to approximately 60% of the initial activity of the freshly purified enzyme.