Rapid method for determination of residual tert-butanol in liposomes using solid-phase microextraction and gas chromatography.

A simple, rapid, and reliable method to detect residual levels of tert-butanol in liposomes using sec-butanol as an internal standard has been developed. Solid-phase microextraction (SPME) followed by gas chromatographic analysis was used to quantify the amount of residual tert-butanol in freeze-dried liposome material. Only 1 min was necessary for reproducible amounts of analyte to absorb onto the SPME fiber, and because this method requires very little sample preparation, a single analysis can be completed in less than 15 min. This method had a linear range of 10-600 microg/mL. Careful control of times of temperature equilibration and exposure to headspace was necessary to ensure reproducible results. This method can easily be applied to other applications in the food and pharmaceutical industries where detection of residual solvents, such as hexane and chloroform, is necessary.

Dissociation of pituitary-adrenal and catecholamine activation after induced cardiac arrest and defibrillation.

To avoid factors which confound attempts to characterize the neuroendocrine response to cardiac arrest, we studied the pituitary-adrenocortical and catecholamine responses to induced ventricular fibrillation (VF) and direct current cardioversion in 10 patients undergoing testing of 'implanted cardioverter defibrillator' devices under sedation. Plasma concentrations of epinephrine were increased 5 min after VF (from a mean basal of 0.39 (S.E.M. 0.09) to a peak of 0.632 (0.212) nmol litre-1; P < 0.05) but were unchanged at other times. Plasma concentrations of norepinephrine did not change at any time. Plasma concentrations of cortisol increased significantly at 10 min (from a mean of 367 (SEM 62) to 539 (64) nmol litre-1; P < 0.001) and remained increased 30 min after VF (470 (74) nmol litre-1; P < 0.05) but had returned towards baseline at 60 min, whereas plasma prolactin concentrations were increased at 5 min (from a mean of 224 (SEM 54) to 320 (63) mu. litre-1; P < 0.01) and remained increased until the end of the sampling period at 60 min (288 (65) mu. litre-1; P < 0.05). Concentrations of adrenocorticotrophic hormone (ACTH) (n = 5) tended to increase but this was not statistically significant. We conclude that a short period of cardiac arrest in lightly sedated humans activated the pituitary-adrenocortical axis but did not appear to stimulate catecholamine secretion. These findings raise questions about the nature and mechanisms of the neuroendocrine response to cardiac arrest.

Dietary menhaden, seal, and corn oils differentially affect lipid and ex vivo eicosanoid and thiobarbituric acid-reactive substances generation in the guinea pig.

This investigation was carried out to characterize the effects of specific dietary marine oils on tissue and plasma fatty acids and their capacity to generate metabolites (prostanoids, lipid peroxides). Young male guinea pigs were fed nonpurified diet (NP), or NP supplemented (10%, w/w) with menhaden fish oil (MO), harp seal oil (SLO), or corn oil (CO, control diet) for 23 to 28 d. Only the plasma showed significant n-3 polyunsaturated fatty acid (PUFA)-induced reductions in triacylglycerol (TAG) or total cholesterol concentration. Proportions of total n-3 PUFA in organs and plasma were elevated significantly in both MO and SLO dietary groups (relative to CO), and in all TAG fractions levels were significantly higher in MO- than SLO-fed animals. The two marine oil groups differed in their patterns of incorporation of eicosapentaenoic acid (EPA). In guinea pigs fed MO, the highest levels of EPA were in the plasma TAG, whereas in SLO-fed animals, maximal incorporation of EPA was in the heart polar lipids (PL). In both marine oil groups, the greatest increases in both docosahexaenoic acid (22:6n-3, DHA) and docosapentaenoic acid (22:5n-3, DPA), relative to the CO group, were in plasma TAG, although the highest proportions of DHA and DPA were in liver PL and heart TAG, respectively. In comparing the MO and SLO groups, the greatest difference in levels of DHA was in heart TAG (MO > SLO, P < 0.005), and in levels of DPA was in heart PL (SLO > MO, P < 0.0001). The only significant reduction in proportions of the major n-6 PUFA, arachidonic acid (AA), was in the heart PL of the SLO group (SLO > MO = CO, P < 0.005). Marine oil feeding altered ex vivo generation of several prostanoid metabolites of AA, significantly decreasing thromboxane A2 synthesis in homogenates of hearts and livers of guinea pigs fed MO and SLO, respectively (P < 0.04 for both, relative to CO). Lipid peroxides were elevated to similar levels in MO- and SLO-fed animals in plasma, liver, and adipose tissue, but not in heart preparations. This study has shown that guinea pigs respond to dietary marine oils with increased organ and plasma n-3 PUFA, and changes in potential synthesis of metabolites. They also appear to respond to n-3 PUFA-enriched diets in a manner that is different from that of rats.

Oxygen transport patterns in patients with sepsis syndrome or septic shock: influence of treatment and relationship to outcome.

OBJECTIVE: To investigate the relationship between oxygen transport patterns and outcome in patients with sepsis syndrome or septic shock managed according to two different treatment regimens. DESIGN: Retrospective study of a subgroup of patients with sepsis syndrome or septic shock taken from a randomized, prospective, controlled trial. SETTING: General intensive care units in a teaching and a district general hospital. PATIENTS: Seventy-eight patients classified according to predetermined criteria as having sepsis syndrome or septic shock were drawn retrospectively from a larger study group of 109 consecutive patients considered to be at risk for developing multiple organ failure. INTERVENTIONS: All patients received volume expansion to an optimal pulmonary artery occlusion pressure. If the therapeutic goals (cardiac index of > 4.5 L/min/m2, oxygen delivery [DO2] of > 600 mL/min/m2, and oxygen consumption [VO2] of > 170 mL/min/m2) were not achieved with fluids alone, patients were randomized to either a control group or a treatment group. In the treatment group, dobutamine (5 to 200 micrograms/kg/min) was administered to increase cardiac index and DO2 until all three goals were simultaneously achieved. In the control group, dobutamine was administered only if the cardiac index was < 2.8 L/min/m2. In both groups, norepinephrine was infused to maintain mean arterial pressure at 80 mm Hg. MEASUREMENTS AND MAIN RESULTS: Hemodynamic, oxygen transport, and lactate measurements were made at the time of admission to the study, at the time of optimal volume administration, at 1, 2, 4, 8, 12, 16, 20, and 24 hrs, then every 6 hrs for the next 24 hrs, and at least every 8 hrs thereafter. The time at which all therapeutic goals were first achieved simultaneously or the time of maximal DO2 was identified and termed "tmax." Survivors from both the control and treatment groups significantly (p < .001) increased cardiac index and DO2 in response to maximal resuscitation, and despite an associated decrease in oxygen extraction (p < .01), there was a significant (p < .01) increase in VO2. In nonsurvivors from both groups, despite significant increases in cardiac index (p < .05) and DO2 (p < .01) at tmax, oxygen extraction decreased (p < .01) and VO2 remained unchanged. DO2 and VO2 were significantly lower (p < .05) at tmax in nonsurvivors than in survivors from both groups. Persistently high lactate concentrations were characteristic of nonsurvivors. CONCLUSIONS: Survivors of sepsis syndrome or septic shock are characterized by an ability to increase both DO2 and VO2. In contrast, nonsurvivors typically have reduced cardiac reserve, they fail to increase VO2 following resuscitation, and when delivery is enhanced with aggressive inotropic support, oxygen extraction falls. These patterns of response were similar in both treatment and control groups, although the magnitude of the changes was exaggerated in the treatment group. These observations may help to explain the findings by some investigators that treatment aimed at achieving survivor values of cardiac index, DO2, and VO2 fails to improve outcome when instituted following admission to intensive care.

Critical illness is associated with low circulating concentrations of insulin-like growth factors-I and -II, alterations in insulin-like growth factor binding proteins, and induction of an insulin-like growth factor binding protein 3 protease.

OBJECTIVES: To describe the sequential changes in the circulating concentrations of insulin-like growth factor-I, insulin-like growth factor-II, and insulin-like growth factor binding proteins in critically ill patients. To determine whether critical illness is associated with induction of a specific protease directed against insulin-like growth factor binding protein 3 and to relate these changes to outcome. DESIGN: Prospective, descriptive study. SETTING: Intensive care unit (ICU) of a university hospital. PATIENTS: Eighteen heterogeneous critically ill patients, requiring ventilatory support. INTERVENTIONS: Serial daily blood samples were collected until death or discharge from the ICU. In five patients, samples were also obtained on the ward before discharge from the hospital. MEASUREMENTS AND MAIN RESULTS: Serum concentrations of insulin-like growth factor-I, insulin-like growth factor-II, and insulin-like growth factor binding proteins 1, 2, and 3 were measured by radioimmunoassay. After 5 days, insulin-like growth factor binding protein 3 concentrations were measured on alternate days. Alterations in binding of insulin-like growth factor-I to insulin-like growth factor binding protein 3 and the presence of protease activity directed against insulin-like growth factor binding protein 3 were investigated by Western ligand blotting. Circulating concentrations of insulin-like growth factor-I and insulin-like growth factor-II were low and remained low throughout the 7-day study period. Insulin-like growth factor binding protein 1 concentrations were initially increased to within the fasting range, but subsequently decreased. There was considerable variability in insulin-like growth factor binding protein 2 concentrations, but generally, concentrations were at the upper end of the normal range throughout. Insulin-like growth factor binding protein 3 concentrations were consistently low and Western ligand blotting at the nadir of the insulin-like growth factor-I concentration demonstrated the presence of a protease directed against insulin-like growth factor binding protein 3. The last recorded concentrations of insulin-like growth factor-I and insulin-like growth factor binding protein 3 were higher in survivors than in nonsurvivors (p < .05). Two patients were also studied for a prolonged period. In one patient, a survivor, insulin-like growth factor-I and insulin-like growth factor binding protein 3 were low initially, but later increased in association with recovery and cessation of protease activity over a period of 33 days. In another patient, a nonsurvivor, insulin-like growth factor-I and insulin-like growth factor binding protein 3 remained low and protease activity persisted until the patient died 38 days after admission to the ICU. CONCLUSIONS: Critical illness is associated with low circulating concentrations of insulin-like growth factor-I, insulin-like growth factor-II, and insulin-like growth factor binding protein 3 and these low values are associated with induction of protease activity specifically directed against insulin-like growth factor binding protein 3. In survivors, recovery is associated with increasing insulin-like growth factor-I and insulin-like growth factor binding protein 3 concentrations and cessation of protease activity. The therapeutic effects of exogenous growth factors are likely to be influenced by these changes.

The differential regulation of the circulating levels of the insulin-like growth factors and their binding proteins (IGFBP) 1, 2 and 3 after elective abdominal surgery.

OBJECTIVES: Patients undergoing abdominal surgery often suffer from morbidity associated with increased protein catabolism. Therapeutic recombinant human insulin-like growth factor (rhIGF)-I has been proposed as a means of reversing this process. As IGFBPs modulate the bioavailability of the IGFs, we have studied the changes in the circulating levels of these peptides during surgery. DESIGN: Patients undergoing elective intestinal surgery were recruited prospectively. Blood samples were taken before, during and after surgery. Standard anaesthetic techniques were used. METHODS: Twelve adults (aged 30-70 years; 9 female, 3 male) undergoing surgery were studied. Serum was taken before premedication (preop), end of surgery (end surg), 2 h, 6 h post surgery, on days 1-4, 7, 10 and 14, and on recovery at 6 weeks. MEASUREMENTS: Serum IGF-I, IGF-II, IGFBP-1, IGFBP-2, IGFBP-3, insulin and C-peptide were measured by radioimmunoassay. IGFBP profiles were also assessed by Western ligand blot (WLB). Samples taken preop and at 2 days were separated by fast-phase liquid chromatography (FPLC) using a Superose 12 column under neutral conditions (pH 7.4), and the fractions were analysed subsequently by WLB and immunoblot using a specific IGFBP-3 antiserum. RESULTS: IGF-I fell rapidly during surgery from 170 +/- 21 (preop) to 133 +/- 14 micrograms/l (end surg) (P < 0.05). The magnitude of this fall could not be explained by haemodilution. IGF-I levels then fell further to a nadir of 103 +/- 10 micrograms/l at day 4 (P < 0.05). IGF-II fell from 580 +/- 46 (preop) to 397 +/- 38 micrograms/l (day 2). Both IGF-I and IGF-II recovered to preop levels at 6 weeks (205 +/- 14 micrograms/l and 623 +/- 30 micrograms/l respectively). IGFBP-3 levels fell similarly from 4.46 +/- 0.45 to 3.2 +/- 0.3 mg/l (end surg) and to a nadir of 2.66 +/- 0.19 mg/l at day 2. There was a close correlation between IGFBP-3 levels and the sum of IGF-I and IGF-II levels before surgery (r = 0.9, P < 0.01) and this was maintained throughout the post-operative period (mean correlation coefficient of 0.86 +/- 0.02, P < 0.05). On days 2 and 3 there was a small but significant increase in the ratio between serum IGF-I and IGFBP-3 levels compared with the preop ratio (P < 0.05 and < 0.005, respectively). WLB demonstrated almost complete absence of IGFBP-3 by day 2. This discrepancy between RIA and WLB analysis of IGFBP-3 suggested the presence of IGFBP-3 protease activity between days 1 and 4. This was confirmed by WLB and immunoblot analyses of samples taken 2 days after surgery. The decrease in IGFBP-3 on WLB was shown to be associated with an increase in the proteolytically cleaved fragments of IGFBP-3. These fragments following FPLC were detected in the high molecular weight fractions, suggesting that the fragments were still able to form the high molecular weight IGFBP-3/ALS complex which is thought to form only when IGF is bound by IGFBP-3. IGFBP-1 levels rose during surgery (mean duration of surgery was 125 minute) from 18 +/- 3 (preop) to 51 +/- 12 micrograms/l (end surg) (P < 0.05). This rise in IGFBP-1 paralleled increases in insulin from 7.3 +/- 1.0 to 20.8 +/- 7.5 mU/l and glucose from 4.6 +/- 0.3 to 8.7 +/- 1.2 mmol/l. IGFBP-1 levels then fell to basal values by 6 hours. IGFBP-2, in contrast, fell slightly during surgery from 636 +/- 14 to 599 +/- 96 mg/l and then returned to basal levels by 6 hours. CONCLUSION: After major surgery there are complex and diverse changes in the IGFs and IGFBPs. The effect of these changes on IGF bioavailability may significantly affect the therapeutic potential of IGF-I in this setting.

Adrenaline administered via a nebulizer in adult patients with upper airway obstruction.

Racemic adrenaline administered via a nebulizer has been used successfully in children with upper airway obstruction resulting from croup and postintubation oedema. We report four adult cases of upper airway obstruction of differing aetiologies successfully managed with the administration of adrenaline via a nebulizer (1 mg in 5 ml of normal saline and repeated as necessary). This appears to be safe and effective in selected cases of upper airway obstruction with immediate benefits and few cardiovascular sequelae.

Elevation of systemic oxygen delivery in the treatment of critically ill patients.

BACKGROUND: Elevation of systemic oxygen delivery and consumption has been associated with an improved outcome in critically ill patients. We conducted a randomized trial to determine whether boosting oxygen delivery by infusing the inotropic agent dobutamine would improve the outcome in a diverse group of such patients. METHODS: On the basis of previously published recommendations, we established the following goals: a cardiac index above 4.5 liters per minute per square meter of body-surface area, oxygen delivery above 600 ml per minute per square meter, and oxygen consumption above 170 ml per minute per square meter. If these goals were not achieved with volume expansion alone, patients were randomly assigned to a treatment or control group. The treatment group received intravenous dobutamine (5 to 200 micrograms per kilogram of body weight per minute) until all three goals had been achieved. Dobutamine was administered to the control group only if the cardiac index was below 2.8 liters per minute per square meter. RESULTS: A total of 109 patients were studied. In nine patients the therapeutic goals were achieved with volume expansion alone; all nine patients survived to leave the hospital. Fifty patients were randomly assigned to the treatment group, and 50 to the control group. During treatment, there were no differences between the two groups in mean arterial pressure or oxygen consumption, despite a significantly higher cardiac index and level of oxygen delivery in the treatment group (P < 0.05). Although the predicted risk of death during hospitalization was 34 percent for both groups, the in-hospital mortality was lower in the control group (34 percent) than in the treatment group (54 percent) (P = 0.04; 95 percent confidence interval, 0.9 to 39.1 percent). CONCLUSIONS: The use of dobutamine to boost the cardiac index and systemic oxygen delivery failed to improve the outcome in this heterogeneous group of critically ill patients. Contrary to what might have been expected, our results suggest that in some cases aggressive efforts to increase oxygen consumption may have been detrimental.

Clinical validation of a radionuclide detector to measure ejection fraction in critically ill patients.

The use of a new non-imaging nuclear probe (Cardioscint) capable of continuous online monitoring of left ventricular function is described in critically ill patients undergoing mechanical ventilation. Ejection fraction, measured by the Cardioscint, was compared with that measured by echocardiography. The mean difference was -1.1% (95% confidence interval -2.9 to +0.6%). Mean difference +/- 2 SD was +10.6 to -12.8% (95% confidence intervals +7.5 to 13.6% and -15.8 to -9.0%, respectively). Examples of fluid loading and inotropic support showed comparable changes in stroke counts measured by the Cardioscint and stroke index measured by thermodilution. The Cardioscint is a practical bedside method for continuous or repeated measurement of ejection fraction and for assessing the response to therapeutic interventions in critically ill patients.

Response of critically ill patients to treatment aimed at achieving supranormal oxygen delivery and consumption. Relationship to outcome.

STUDY OBJECTIVE: To evaluate the response to therapy aimed at achieving supranormal cardiac and oxygen transport variables (cardiac index [CI] > 4.5 L/min/m2, oxygen delivery [DO2] > 600 ml/min/m2, and oxygen consumption [VO2] > 170 ml/min/m2) in a heterogenous group of critically ill patients and to assess its relationship to outcome. DESIGN: Patients were divided retrospectively into two groups. Group 1 (n = 15) achieved supranormal values for CI, DO2 and VO2 simultaneously during the first 24 h. Group 2 (n = 17) failed to achieve these goals simultaneously at any time point. SETTING: General intensive care units in a teaching and a district general hospital. PATIENTS: Thirty-two patients at risk of developing multiple organ failure were studied prospectively. INTERVENTIONS: Patients received volume expansion and then, if necessary, dobutamine (5 to 200 micrograms/kg/min) to increase CI and DO2 until all three goals were achieved simultaneously. RESULTS: In group 2, target VO2 could never be reached despite the fact that 11 (65 percent) patients achieved target CI and DO2 simultaneously. In this group, lactate levels did not fall and 16 patients died. In contrast, in group 1, attainment of all goals was associated with a significant reduction (p < 0.05) in blood lactate levels, and all but one of these patients survived. The persistently raised lactate levels in group 2 were associated with significantly higher venous oxygen saturation (SvO2) and lower oxygen extraction ratio (OER); in these patients, SvO2 rose and OER fell in response to increases in DO2. CONCLUSION: These results suggest that failure to increase VO2 was related predominantly to an inability of the tissues to extract or utilize oxygen rather than a failure to increase DO2. These findings support the hypothesis that in order to survive a critical illness, patients must achieve a high level of VO2. An inability to do so is reflected in persistently elevated blood lactate levels and an extremely poor prognosis.

The relationship between cardiac reserve and survival in critically ill patients receiving treatment aimed at achieving supranormal oxygen delivery and consumption.

The relationship between survival and cardiac responsiveness to therapy aimed at achieving supranormal values for cardiac index, oxygen delivery and oxygen consumption (cardiac index (CI) > 4.5 l/min/m2, oxygen delivery (DO2) > 600 ml/min/m2, and oxygen consumption (VO2) > 170 ml/min/m2), has been investigated in a heterogeneous group of critically ill patients. Thirty-two patients were prospectively studied and divided into survivors and non-survivors. Cardiac reserve was assessed by determining changes in CI, left ventricular stroke work index (LVSWI) and cardiac power output (CPO) in response to optimal fluid administration and inotropic stimulation with dobutamine. On admission LVSWI and CPO were significantly higher in survivors (P < 0.05), despite no significant differences in pulmonary artery occlusion pressure (PAOP). In response to fluid CI, CPO and LVSWI increased significantly in survivors (P < 0.01), but not in non-survivors. Following optimal fluid administration, survivors achieved significantly higher values for CI, LVSWI (P < 0.01), and CPO (P < 0.001) than non-survivors. At maximum resuscitation all three variables were significantly higher in survivors than in non-survivors (P < 0.001). The dose of dobutamine administered to non-survivors (median (range) 100 (5-200)) was significantly greater (P < 0.001) than that given to the survivors (median (range) 10 (0-25)). The dose of dobutamine was limited by complications in 12 of the non-survivors. These observations suggest that cardiac reserve is an important determinant of outcome following critical illness. In unresponsive patients attempts to achieve supranormal oxygen delivery and consumption with massive inotropic support may not only be ineffective but frequently precipitates tachydysrhythmias and myocardial ischaemia.

Bone marrow hypervascularity in patients with myelofibrosis identified by infra-red thermography.

Infra-red thermography was used to assess bone marrow vascularity in six patients with myelofibrosis secondary to myeloproliferative disorders (four primary myelofibrosis and two primary proliferative polycythaemia). The technique was evaluated with conventional static and dynamic radio-isotopic imaging and with immunohistochemical staining of bone marrow biopsies. Infra-red thermography identified increased bone marrow blood flow in patients with established myelofibrosis and correlated with dynamic radio-isotopic studies of blood flow and hypervascularity identified by immunohistochemistry. Increased bone marrow blood flow and vascular proliferation was not confined to the central bone marrow but also extended into the peripheral marrow of the long bones. Endothelial cell proliferation may be an initiating event in the pathogenesis of myelofibrosis but evaluation of bone marrow vascularity and blood flow has hitherto relied on invasive and complicated techniques. This study has identified bone marrow hypervascularity in patients with myelofibrosis and shown infra-red thermography to be a simple non-invasive method of assessing vascularity. This non-invasive technique may be used to study disease progression and response to therapeutic regimens in patients with myelofibrosis and to study bone marrow blood flow in other bone marrow disorders.

Survival in adults after cardiac arrest due to drowning.

Some remarkable cases of full neurological recovery after cardiac arrest following immersion incidents have been intermittently reported in the journals over the years. These have largely been in children or teenagers who have fallen into extremely cold water. We report here two older adults who recovered completely after a period of cardiac arrest in cold water. Certainly, death should not be pronounced in cold water drowning, without a thermometer reading and ECG.

Argyria or cyanosis.

A case of argyria secondary to the prolonged use of nasal drops that contained mild silver protein is presented. The patient was thought to be cyanosed immediately after a total knee replacement. Argyria, its differentiation from cyanosis and its prevention are discussed. The need for a careful drug history is noted.

Gastric emptying: a study to compare the effects of intrathecal morphine and i.m. papaveretum analgesia.

Gastric emptying was studied in two groups of 10 patients who underwent elective cholecystectomy. The groups were comparable for age, weight and duration of operation. Gastric emptying was measured with a radioisotopic technique using Tc99m-DTPA (diethylene triamine pentaacetic acid) before, and 24 h after, surgery. Analgesia was provided by intrathecal morphine 0.8 mg alone (group A) or by i.m. papaveretum 10 mg, administered as required, plus one additional dose 1 h before the postoperative measurement (group B). Control gastric emptying rates were not significantly different in the two groups (mean +/- SD: A = 76.6 +/- 23.0 ml; B = 81.8 +/- 16.3 ml in 30 min). After surgery, gastric emptying was significantly greater in group A (42.9 +/- 35.6 ml) than in group B (11.0 +/- 27.9 ml) (P less than 0.05).

Mass spectra of fatty acid derivatives, of isopropylidenes of novel glyceryl ethers of cod muscle and of phenolic acetates obtained with the Finnigan MAT Ion Trap Detector.

The applicability of the Finnigan MAT Ion Trap Detector (ITD) mass spectrometer for structure determination in some selected fatty acids and their derivatives has been investigated. Isopropylidene derivatives of novel glyceryl ethers isolated from cod flesh and of phenolic acetates are included to indicate the potential for diverse structures and to clarify the protonation of ions. The ITD is a simple and unsophisticated gas liquid chromatograph-mass spectrometer, but the spectra obtained are in most respects comparable to those from more conventional electron impact mass spectrometers. However, due to the comparatively high background pressure (approximately 10(-3) torr) in the ionization chamber of the ITD, there is a tendency for both neutral and ionized molecules to acquire protons from other molecules or fragments through collision. In many cases, the molecular ion was observed as the protonated molecular ion (M + 1), as in chemical ionization mass spectrometry. These interactions can be minimized if the sample load is decreased. Phenolic acetates exhibit not only protonation of the molecular ion, but also protonation of stable fragmented neutral molecules or ions.