Collaborative study for the calibration of the Ph. Eur. prekallikrein activator in albumin BRP batches 4, 5 and 6.

An international collaborative study was organised by the European Directorate for the Quality of Medicines & HealthCare (EDQM, Council of Europe) to calibrate replacement batches for the current European Pharmacopoeia (Ph. Eur.) prekallikrein activator (PKA) in albumin biological reference preparation (BRP), whose stocks were dwindling. The study was run in the framework of the Biological Standardisation Programme (BSP) of the Council of Europe and the European Union (EU) Commission. Twenty three laboratories from official medicines control authorities and manufacturers in Europe and outside Europe took part in the study. Three candidate replacement batches were produced from the same material as the one used for the World Health Organization (WHO) 2(nd) International Standard (IS) for PKA in albumin (02/168) and the Ph. Eur. PKA in albumin BRP batches 1, 2 and 3. Participants were requested to evaluate the candidate batches against the current WHO IS using their routine assay method. The Ph. Eur. PKA in albumin BRP batch 3 (BRP3) was also included in the test panel to ensure the continuity of the consecutive BRP batches. The study confirmed the stability of the PKA content of the current BRP3. The candidate batches were found to be comparable. Previous data on the starting material support its high stability. Thermal stress study on the candidate batches confirmed the stability of their PKA activity. The Commission of the Ph. Eur. officially adopted in November 2013 the 3 candidate batches as Ph. Eur. PKA in albumin BRP batches 4, 5 and 6 with an assigned content of 38 IU/vial. The activity of the 3 new batches of Ph. Eur. PKA in albumin BRP will be regularly monitored.

Spectroscopy of nS, nP, and nD Rydberg series of Cs atoms on helium nanodroplets.

The preparation of an artificial superatom consisting of a positive charge inside a superfluid helium nanodroplet and an electron in an orbital surrounding the droplet is of fundamental interest and represents an experimental challenge. In this work, nanodroplets of several thousand helium atoms are doped with single caesium (Cs) atoms. While on the droplet, the Cs valence electron is excited in two steps through an intermediate state into nS, nP, and nD states. The excitation is monitored by laser induced fluorescence or, for high principal quantum numbers, by resonant three-photon-ionization. On-droplet Rydberg excitations are resolved up to about n = 20. The energies are compared with those of free Cs atom Rydberg states and quantum defects as well as the on-droplet ionization threshold are derived.

Establishment of replacement batches for prekallikrein activator in albumin biological reference preparation.

A collaborative study was run by the European Directorate for the Quality of Medicines and HealthCare (EDQM) under the aegis of the Biological Standardisation Programme (BSP) to establish replacement batches of the current Prekallikrein activator in albumin Biological Reference Preparation (BRP) batch 1, the stocks of which were dwindling. Candidate BRP replacement batch 2 and batch 3 were assayed against the 2nd World Health Organization International Standard for Prekallikrein activator, human (2nd IS) and the Prekallikrein activator in albumin BRP batch 1. The candidate batches were manufactured from the same starting material as the current Biological Reference Preparation and the 2nd IS. They consisted of a 20 % solution of albumin lyophilised under the same conditions as the Prekallikrein activator in albumin BRP batch 1. Sixteen laboratories participated in the collaborative study and were requested to assay the candidates by their routine method, complying with the European Pharmacopoeia (Ph. Eur.) general method 2.6.15 for the determination of prekallikrein activator content. A central statistical analysis was performed at the EDQM using in-house calculations of prekallikrein activator contents provided by the participating laboratories. On the basis of the results of this study, which confirmed the assigned potency of 29 IU/vial of Prekallikrein activator in albumin BRP batch 1, the 2 candidate materials were assigned a potency of 30 IU/vial. The 2 candidates were adopted by the Ph. Eur. Commission in March 2008 as Ph. Eur. Prekallikrein activator in albumin Biological Reference Preparation batch 2 and batch 3.

Collaborative study to establish a new biological reference preparation for prekallikrein activator.

An International Collaborative Study was organized to replace the current World Health Organization (WHO) International Standard (IS) for Prekallikrein Activator (PKA) and to establish a European Pharmacopoeia (Ph. Eur.) Biological Reference Preparation (BRP). The project was jointly organized by the European Directorate for the Quality of Medicines (EDQM) and the National Institute for Biological Standards and Control (NIBSC) to identify and calibrate suitable materials that could act as an IS and a Ph. Eur. BRP. The current IS for PKA (82/530) is popular and stocks are declining rapidly, therefore necessitating calibration of a replacement. A Ph. Eur. BRP is needed, as PKA control on the finished product is part of the Official Control Authority Batch Release (OCABR) of Human Albumin. The current IS, 82/530 is a 5 per cent albumin solution spiked with purified PKA. However, during planning stages it was decided that the replacement IS (and BRP) should be made from a 20 per cent albumin preparation containing a significant level of PKA as the current IS is used to measure PKA in albumin and high levels are more likely to be encountered in more concentrated 20 per cent solutions. A suitable material was sourced by the EDQM and filled into ampoules at NIBSC and vials by the EDQM. Both preparations were included in the collaborative study that involved 31 laboratories from 17 countries. Another important goal of this study was to investigate the influence of the prekallikrein substrate (PKS) on PKA determination in albumin solutions following earlier concerns that variability amongst PKS prepared in-house could significantly affect PKA determinations. Laboratories were requested to perform their routine assays following Ph. Eur. guidelines and recommendations on doses, replication and randomization were also provided to study participants. Participants were requested to use material A (the current IS, 82/530) to perform at least 4 assays to determine PKA levels in sample B (NIBSC ampouled material, candidate IS, 02/168), sample C (EDQM material in vials candidate Ph. Eur. BRP Batch 1), and sample D (an ampouled preparation of 2.5 per cent albumin containing a lower level of PKA). A commercial substrate was provided for participants to perform half the assays and the remaining assays were to be performed using the laboratories' in-house substrate (where available). Collation of participants' results showed that samples B and C had the same level of PKA of 29 IU/ampoule, the concentration anticipated from development studies. Importantly, there was no significant difference between the PKA level obtained using the commercial substrate provided and the laboratories' own in-house substrate. Previous observations on lyophilized preparations of PKA indicate that the enzyme is very stable. Detailed investigations conducted in this study show that the PKA in albumin used to make samples B and C is very stable and suitable for long-term storage as a reference material.

An international collaborative study to replace the 1st international standard for prekallikrein activator.

BACKGROUND AND OBJECTIVES: The aim of this study was to replace the 1st International Standard (IS) for prekallikrein activator (PKA) (code 82/530) with a new IS and European Biological Reference Preparation (BRP). The new standards were freeze dried 20% albumin solution containing PKA, the same solution that is tested using these standards. Aspects of the methodology for PKA determination were also examined as part of this study. MATERIALS AND METHODS: A batch of 20% albumin containing approximately 30 IU/ml was donated by a manufacturer of blood products and dispensed into ampoules at the National Institute for Biological Standards and Control (NIBSC) to create the candidate IS (02/168, sample B in the study) and at the European Directorate for the Quality of Medicines (EDQM) to create the candidate BRP (sample C in the study). The concentration of PKA in these preparations was determined in an international collaborative study involving 31 laboratories from 17 countries worldwide in comparison with the 1st IS for PKA (82/530) containing 85 IU of PKA per ampoule. Participants were requested to perform their own in-house method, based on the current Ph. Eur. monograph for determination of PKA in albumin solutions. Participants were provided with sufficient samples to perform two or three assays and were asked to use their local prekallikrein substrate (PKS) and also to use a commercial PKS provided as part of the study, in order to investigate the importance of the source of PKS on the final potency values. RESULTS: Samples B and C emerged with identical PKA concentrations of 29 IU/ml, very close to the expected value. This figure was determined using a variety of statistical methods, with the participants' own calculated values and values calculated centrally at the EDQM using raw data. The value of 29 IU/ml was consistent and independent of the method of calculation, although interlaboratory variability was more sensitive to the statistical analysis method. There was no statistically significant difference in mean potencies when comparing results with the laboratories' own local substrate and the substrate provided for the study. All stability studies indicate that these lyophilized preparations of PKA in 20% albumin are extremely stable. CONCLUSIONS: Samples B and C were established as the 2nd IS (code 02/168) and PKA activator in albumin BRP batch 1 (Y0000263), respectively, with a potency of 29 IU per ampoule. Results from this study indicate that testing for PKA in albumin may be less sensitive to the source of PKS than previously feared. The study highlights a number of methodological issues that may need revising in the Ph. Eur. general chapter 2.6.15.

Intraoperative catecholamine release in brain-dead organ donors is not suppressed by administration of fentanyl.

BACKGROUND AND OBJECTIVE: Endogenous catecholamines are released in brain-dead organ donors following painful stimulation during retrieval surgery, and might be harmful to harvested organs. Our hypothesis was that inhibition of pain by fentanyl would inhibit such catecholamine release. METHODS: We tested 17 brain-dead organ donors in a randomized, placebo-controlled, double-blinded study. Blood samples for determination of epinephrine and norepinephrine concentrations were obtained before and 10 min after in take of either fentanyl 7 microg kg(-1) or an equivalent volume of placebo. Further points of measurement were taken after skin incision and sternotomy. Mean arterial pressure and heart rate at these points were recorded. RESULTS: Catecholamine concentrations rose following painful stimuli. No differences in haemodynamics, between the fentanyl and the placebo group were detectable. Epinephrine concentrations, but not those of norepinephrine, were higher in the fentanyl group, reaching significance following sternotomy. CONCLUSION: We conclude that the use of fentanyl (7 microg kg(-1)) was not effective in suppressing the catecholamine release, following painful surgical stimulation in brain-dead organ donors.

Recollection of dreams after short general anaesthesia: influence on patient anxiety and satisfaction.

BACKGROUND AND OBJECTIVE: We ascertained whether dreams during short general anaesthesia influence subsequent patient satisfaction and anxiety. METHODS: Fifty female patients were randomized into two groups to test for a difference between intravenous and inhalational anaesthesias. In Group Propo, anaesthesia was induced and maintained with propofol; in Group Metho-Iso, anaesthesia was induced with methohexital and maintained with isoflurane. Satisfaction and anxiety with anaesthesia were evaluated using a visual analogue scale from 0 to 100. Dream incidence rate, satisfaction and anxiety were assessed from immediately after waking until 3 months later. RESULTS: Seventeen patients (34%) dreamed during anaesthesia. There were no significant differences in satisfaction or anxiety after anaesthesia between the dreaming and non-dreaming patients (satisfaction, 92.3 +/- 21.6 versus 92.1 +/- 21.6; anxiety, 21.1 +/- 21.1 versus 30.3 +/- 32.1), or between Group Propo and Group Metho-Iso (satisfaction, 94.4 +/- 19.3 versus 90.0 +/- 23.4; anxiety, 26.0 +/- 27.6 versus 28.4 +/- 30.7). There was no significant difference in the incidence rate of dreaming with the type of anaesthesia used (Group Propo, 11 patients; Group Metho-Iso, 6 patients). CONCLUSIONS: Dreaming during general anaesthesia is common but does not influence satisfaction or anxiety after anaesthesia.

Comparable postoperative pulmonary atelectasis in patients given 30% or 80% oxygen during and 2 hours after colon resection.

BACKGROUND: High concentrations of inspired oxygen are associated with pulmonary atelectasis but also provide recognized advantages. Consequently, the appropriate inspired oxygen concentration for general surgical use remains controversial. The authors tested the hypothesis that atelectasis and pulmonary dysfunction on the first postoperative day are comparable in patients given 30% or 80% perioperative oxygen. METHODS: Thirty patients aged 18-65 yr were anesthetized with isoflurane and randomly assigned to 30% or 80% oxygen during and for 2 h after colon resection. Chest radiographs and pulmonary function tests (forced vital capacity and forced expiratory volume) were obtained preoperatively and on the first postoperative day. Arterial blood gas measurements were obtained intraoperatively, after 2 h of recovery, and on the first postoperative day. Computed tomography scans of the chest were also obtained on the first postoperative day. RESULTS: Postoperative pulmonary mechanical function was significantly reduced compared with preoperative values, but there was no difference between the groups at either time. Arterial gas partial pressures and the alveolar-arterial oxygen difference were also comparable in the two groups. All preoperative chest radiographs were normal. Postoperative radiographs showed atelectasis in 36% of the patients in the 30%-oxygen group and in 44% of those in the 80%-oxygen group. Relatively small amounts of pulmonary atelectasis (expressed as a percentage of total lung volume) were observed on the computed tomography scans, and the percentages (mean +/- SD) did not differ significantly in the patients given 30% oxygen (2.5% +/- 3.2%) or 80% oxygen (3.0% +/- 1.8%). These data provided a 99% chance of detecting a 2% difference in atelectasis volume at an alpha level of 0.05. CONCLUSIONS: Lung volumes, the incidence and severity of atelectasis, and alveolar gas exchange were comparable in patients given 30% and 80% perioperative oxygen. The authors conclude that administration of 80% oxygen in the perioperative period does not worsen lung function. Therefore, patients who may benefit from generous oxygen partial pressures should not be denied supplemental perioperative oxygen for fear of causing atelectasis.

Postoperative pain and subcutaneous oxygen tension.

Surgical patients randomly assigned to standard pain control had postoperative subcutaneous oxygen partial pressures that were significantly less than patients given better pain treatment. Our data suggest that control of postoperative pain is a major determinant of surgical-wound infection and should be given the same consideration as maintaining adequate vascular volume and normothermia.

[Neostigmine and edrophonium. Antagonism of profound and shallow mivacurium blockade]. / Neostigmin und Edrophonium. Antagonisierung einer tiefen und oberflächlichen Mivacuriumblockade.

UNLABELLED: Mivacurium has a short duration of action because it is rapidly hydrolysed by plasma cholinesterase. There is ongoing controversy concerning the antagonism of mivacurium-induced neuromuscular block, firstly because of its short spontaneous recovery time, and secondly because the metabolism of mivacurium may be inhibited by anticholinesterases. We therefore compared neostigmine and edrophonium reversal of deep and moderate mivacurium-induced blocks. METHODS: After approval by the local ethics committee, 48 ASA class I and II adult patients were investigated during nitrous oxide-fentanyl-thiopental anaesthesia using train-of-four (TOF) stimulation and monitoring of the isometric force of adduction of a thumb. The patients received 0.2 mg/kg mivacurium i.v. Neuromuscular transmission was allowed to recover spontaneously in 10 patients (group SP). In 2 other groups the neuromuscular block was antagonised by administration of 0.04 mg/kg neostigmine (group N5; n = 9) or 1.0 mg/kg edrophonium (group E5; n = 10) when T1 had recovered spontaneously to 5% of control. In two other groups the neuromuscular block was antagonised with the same doses of neostigmine or edrophonium in 10 patients (group N25) and 9 patients (group E25), respectively, when T1 had recovered spontaneously to 25% of control. RESULTS: Neostigmine or edrophonium administered when T1 had recovered spontaneously to 25% of control shortened the recovery time (time from administration of ant-agonist to a T4/T1-ratio of 0.7) significantly from 10.7 +/- 2.2 min (mean +/- SD) in the SP group to 5.1 +/- 2.0 and 5.3 +/- 1.5 min in the N25 and E25 groups, respectively (P < 0.05). The corresponding recovery times in the SP, N5, and E5 groups were 15.9 +/- 2.9, 10.0 +/- 1.9, and 7.7 +/- 2.2 min, respectively. The difference between the SP and E5 groups was significant (P < 0.05). The recovery indices (time from 25% to 75% recovery of T1) of 3.0 +/- 1.3 and 1.7 +/- 0.9 min for the E5 and E25 groups, respectively, were shorter than those of the SP group at 6.1 +/- 2.0 min (P < 0.05). CONCLUSIONS: Two theoretical reasons, the very rapid onset time and the fact that it does not inhibit plasma cholinesterase, suggest edrophonium to be the preferred antagonist of a mivacurium-induced blockade. These two characteristics are reflected in our results: only edrophonium was able to shorten the recovery index significantly and, administered at a profound level of mivacurium-induced neuromuscular block, only edrophonium was successful in shortening recovery time significantly. Therefore, edrophonium should be the anticholinesterase of choice to antagonise a mivacurium-induced neuromuscular block.

Mild intraoperative hypothermia prolongs postanesthetic recovery.

BACKGROUND: Intraoperative hypothermia is common and persists for several hours after surgery. Hypothermia may prolong immediate recovery by augmenting anesthetic potency, delaying drug metabolism, producing hemodynamic instability, or depressing cognitive function. Accordingly, the authors tested the hypothesis that intraoperative hypothermia prolongs postoperative recovery. METHODS: Patients undergoing elective major abdominal surgery (n = 150) were anesthetized with isoflurane, nitrous oxide, and fentanyl. They were randomly assigned to routine thermal management (hypothermia) or extra warming (normothermia). Postoperative surgical pain was treated with patient-controlled analgesia. Fitness for discharge from the postanesthesia care unit was evaluated at 20-min intervals by investigators blinded to group assignment and postoperative core temperatures. Scoring was based on a modification of a previously published system that included activity, ventilation, consciousness, and hemodynamic responses. Patients were considered fit for discharge when they sustained a score of 80% (13 points) for at least two consecutive measurement periods. RESULTS: Morphometric characteristics and anesthetic management were similar in each group. Final intraoperative core temperatures differed by approximately 2 degrees C: 34.8 +/- 0.6 versus 36.7 +/- 0.6 degrees C (mean +/- SD, P < 0.001). Postoperative pain scores and postoperative use of patient-controlled opioid were similar. Hypothermic patients required approximately 40 min longer (94 +/- 65 vs. 53 +/- 36 min) to reach fitness for discharge, even when return to normothermia was not a criterion (P < 0.001). Duration of recovery in the two groups differed by approximately 90 min when a core temperature >36 degrees C was also required (P < 0.001). CONCLUSION: Maintaining core normothermia decreases the duration of postanesthetic recovery and may, therefore, reduce costs of care.

Influence of prostaglandin E1 on tissue ischemia during surgical repair of the abdominal aorta.

OBJECTIVE: To evaluate tissue protection by PGE1 during leg ischemia in patients undergoing aortic surgery. DESIGN: Randomized, controlled prospective clinical trial. SETTING: Single university hospital. PARTICIPANTS: 19 consecutive patients undergoing abdominal aortic aneurysm repair. INTERVENTIONS: Patients received infusions of 30 ng/kg/min of PGE1 or saline. MEASUREMENTS AND MAIN RESULTS: Hemodynamic variables, lactate, creatine phosphokinase, and thromboxane B2 (TXB2) were measured. In the control group, the decrease in cardiac index (CI) after aortic cross-clamping (AXC) persisted until unclamping together with a decrease in femoral venous O2 content (CfvO2). In the PGE1 group, CI returned to baseline with a trend toward greater CfvO2 levels. During reperfusion in the PGE1 group, O2 consumption and lactate levels exceeded preclamp values. Pulmonary hypertension occurred equally in both groups but did not correlate with TXB2, which was not altered by surgery or by PGE1 infusion. CONCLUSIONS: Intraoperative PGE1 treatment offers no benefit and may exacerbate tissue ischemia during AXC by redistributing microcirculatory flow or limiting cellular oxygen utilization in a manner that overwhelms any possible protective effect.

Endocrine stress reaction to surgery in brain-dead organ donors.

We studied the course of plasma levels of the stress markers adrenocorticotropic hormone (ACTH), cortisol, human growth hormone (h-GH), beta-endorphin, and prolactin during retrieval surgery in eleven brain-dead organ donors scheduled for multiple organ explantation. Donors were divided into two groups according to hemodynamic stability. Hormones demonstrated a great variability in plasma levels and in the pattern of reaction, revealing a different degree of remaining pituitary function. Beta-Endorphin was the only stress hormone that showed a response to surgical stimuli in six patients. Only three of them developed a concomitant rise in ACTH. Cortisol, prolactin, and h-GH plasma levels did not change during the observation period. In the three cases with a slight elevation in ACTH, no subsequent change in cortisol was detectable. Beta-Endorphin showed greater variability and a tendency to higher levels in the group presenting with a higher arterial pressure, which resulted in a significant difference (P < 0.005) when distributions were compared using the Mann-Whitney U-test. No correlation was found between hypotensive episodes and deficiencies of other stress hormones. We conclude that pituitary function varies considerably in brain-dead organ donors without demonstrating a correlation to the onset of hypotension. Thus, we feel no need for a substitution treatment with any of the hormones investigated prior to organ explanation.

Postoperative hemodynamic and thermoregulatory consequences of intraoperative core hypothermia.

STUDY OBJECTIVE: To evaluate the postoperative hemodynamic and thermoregulatory consequences of intraoperative core hypothermia. DESIGN: Prospective, randomized clinical trial. SETTING: Operating room and postanesthesia care unit of a university hospital. PATIENTS: 74 healthy, ASA status I, II, and III patients (average age 58 yrs) undergoing elective colon surgery. INTERVENTIONS: Patients were randomly assigned to be kept normothermic or approximately 2.5 degrees C hypothermic during surgery. Anesthesia was maintained with isoflurane, nitrous oxide, and fentanyl. Postoperatively, surgical pain was treated with patient-controlled analgesia (PCA) opioid. MEASUREMENTS AND MAIN RESULTS: An observer blinded to group assignment and core temperatures evaluated shivering, thermal comfort, surgical pain, heart rates (HRs), and blood pressures (BPs) during the first six postoperative hours. Morphometric characteristics, oxygen saturation, fluid balance, PCA-administered opioid, and visual analog pain scores were comparable in the two groups. Hypothermic patients felt uncomfortably cold during recovery, and their postoperative core temperatures remained significantly less than in the normothermic patients for more than four hours. Peripheral vasoconstriction and shivering were common in the hypothermic patients but rare in those kept normothermic. HRs and BPs were comparable in the two groups. CONCLUSIONS: These data confirm that the effects of intraoperative hypothermia on postoperative HR and BP are modest in relatively young, generally healthy patients. In contrast, intraoperative hypothermia caused substantial postoperative thermal discomfort, and full recovery from hypothermia required many hours. Delayed return to care normothermia apparently resulted largely from postoperative thermoregulatory impairment.

The influence of proteins on silver staining of nucleic acids following polyacrylamide gel electrophoresis.

The influence, on silver staining, of proteins and restriction enzymes in polymerase chain reaction (PCR) products was studied in 12-20% polyacrylamide gels. For small DNA fragments (74, 41 and 33 bp) best results were achieved using 20% polyacrylamide gels. In 12% gels, restriction enzymes and also proteins used for PCR (bovine serum albumin, Taq-DNA polymerase) interfere with silver staining of nucleic acids.

Cardiovascular and catecholamine response to surgery in brain-dead organ donors.

Eleven brain-dead organ donors were studied during surgery. Plasma levels of adrenaline and noradrenaline were measured before and after skin incision, upon sternotomy and 15, 30 and 45 min thereafter. Haemodynamic changes were measured continuously throughout the observation period. Blood pressure and heart rate increased after skin incision, remained high at sternotomy then decreased towards the end of the observation period in six of the 11 patients. Plasma catecholamines increased promptly with the onset of surgical stimuli. We conclude that surgical stress can evoke an excessive rise of plasma adrenaline and noradrenaline and thus could impair allograft function.

Pharmacodynamics, pharmacokinetics, and intubation conditions after priming with three different doses of vecuronium.

The effects of three different priming doses of vecuronium on pharmacokinetics, pharmacodynamics, and endotracheal intubation conditions were investigated. Forty-two patients were studied in two parts. In each part, 21 patients were allocated into three groups (n = 7/group) receiving 10, 15, or 20 micrograms/kg vecuronium as a priming dose, followed by a 50- micrograms/kg intubating dose 6 min later. In Part I, Train-of-Four (TOF) ratios and serum concentrations after priming were measured every minute up to the sixth minute. Onset time [from injection of the intubating dose to maximum depression of the first twitch (T1)], clinical duration (T1 return from maximum block to 25% of control), and recovery index (T1 recovery from 25% to 75% of control) were calculated and serum concentrations were determined up to 6 h after injection of the intubating dose. In Part II, the intubating dose was injected 4 min after priming, onset time was determined, and intubation conditions were scored. TOF ratio was significantly lower after priming with 20 micrograms/kg at the fifth and sixth minutes (0.59 +/- 0.29 and 0.56 +/- 0.32; mean +/- 1 SD) compared with the first minute (0.95 +/- 0.1). Recovery index was significantly increased after priming with 20 micrograms/kg (13.2 +/- 6.6 min, P < 0.05) compared with 10 micrograms/kg (9.2 +/- 4.8 min) and 15 micrograms/kg (6.7 +/- 1.5 min). Between groups no differences in onset time, clinical duration, and pharmacokinetic variables were found. In Part II, onset time and intubating scores showed no significant differences between the groups.(ABSTRACT TRUNCATED AT 250 WORDS)