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Development of a successful research program can seem daunting when looked at from the starting line. It will take years if not decades to succeed and become sustainable. It requires local partnerships and mentoring; it mandates the establishment of review boards; it requires national health policies to allow for protected time for research in salaries and for fund granting agencies to be set up; it requires training of researchers and support staff as well as a change in the mindset of clinical staff on the floor. It will almost inevitably require international support of some kind for low- and middle-income country researchers, be it university programs or other academic or private institutions. Success can occur; most likely it will occur by partnering with local research experts outside of emergency medicine in some combination with international networks and mentoring. Perhaps the most critical elements to success are intellectual curiosity and a burning flame of passion - and neither of those carry a financial cost., Competing Interests: The authors declared no conflicts of interest., (© 2020 African Federation for Emergency Medicine. Publishing services provided by Elsevier.)

Bioconjugation, defined as chemical modification of biomolecules, is widely employed in biological and biophysical studies. It can expand functional diversity and enable applications ranging from biocatalysis, biosensing and even therapy. This review summarizes how chemical modifications of cytochrome P450 enzymes (P450s or CYPs) have contributed to improving our understanding of these enzymes. Genetic modifications of P450s have also proven very useful but are not covered in this review. Bioconjugation has served to gain structural information and investigate the mechanism of P450s via photoaffinity labeling, mechanism-based inhibition (MBI) and fluorescence studies. P450 surface acetylation and protein cross-linking have contributed to the investigation of protein complexes formation involving P450 and its redox partner or other P450 enzymes. Finally, covalent immobilization on polymer surfaces or electrodes has benefited the areas of biocatalysis and biosensor design. This article is part of a Special Issue entitled: Cytochrome P450 biodiversity and biotechnology, edited by Erika Plettner, Gianfranco Gilardi, Luet Wong, Vlada Urlacher, Jared Goldstone., (Copyright © 2017 Elsevier B.V. All rights reserved.)

A rapid and simple method for the determination of morphine (M), normorphine (NM), morphine-3-glucuronide (M3G) and morphine-6-glucuronide (M6G) in plasma by high-performance liquid chromatographic separation with mass spectrometric detection (HPLC-MS) has been developed. Samples (40 microl) were cleaned-up by protein precipitation with two volumes (80 microl) of acetonitrile and reconstituted in formic acid 0.1% in water. Naloxone was used as internal standard. Analytes were separated on a phenyl-hexyl column using a step-gradient (1 ml/min) of acetonitrile and formic acid in water. Acetonitrile was added post-column (0.3 ml/min). Quantification of morphine and its metabolites was achieved with an Agilent 1100 series HPLC-MS system equipped with electrospray interface set to selected ion-monitoring (SIM) mode. Calibration curves covered a wide range of concentrations (2.44-10,000 nM) and were best fitted with a weighed quadratic equation. The limits of quantification achieved with this method were 2.44 nM for M and 4.88 nM for NM, M3G and M6G. The method proved accurate (85-98%), precise (C.V.<10%) and was successfully applied to a wide range of in vitro and in vivo pharmacokinetic studies in rodents.

Many opioids are substrates of the efflux transporter P-glycoprotein (P-gp) in the blood-brain barrier (BBB). In situ brain perfusion in wild-type and mdr 1a(-/-) P-gp-deficient mice was utilized to investigate potential P-gp-mediated transport of novel nonpeptidic delta agonists (AR-M delta compounds). Because radioactive compounds were not available for this series, liquid chromatography-mass spectrometric detection (LC-MS) was the assay methodology of choice. Verapamil in the perfusion buffer (0.5 microM) served as a positive control for P-gp-mediated efflux and as an experimental internal standard for P-gp modulation by AR-M delta compounds. LC-MS provided excellent assay sensitivity with no significant interferences. In P-gp-competent mice, the brain extraction of AR-M delta compounds ranged from 1.1 to 96%. The ratio of initial brain uptake clearances (Cl(up)) in P-gp-deficient and wild-type mice (P-gp effect) ranged from 0.96 to 4.91. Some compounds increased the Cl(up) of verapamil in P-gp-competent mice, consistent with P-gp inhibition. These results demonstrate that LC-MS is an appropriate assay methodology for mouse brain perfusion samples, that AR-M delta compounds may interact with P-gp in the BBB, and that the internal strategy can provide useful information concerning P-gp modulation by compounds of interest., (Copyright 2002 Wiley-Liss, Inc.)

Objectives: Errorless compliance training is a success-based, noncoercive intervention for children with severe oppositional behavior. The strategy involves hierarchical introduction of more demanding parental requests at a gradual pace that greatly reduces noncompliance and obviates the need for constraining consequences (e.g., time-out). In this study, this approach was evaluated as a treatment for severe disciplinary problems in children from violent homes., Method: Participants were 15 children (aged 3-10 years) and their mothers. All participants had experienced long-term family violence. Using a multiple-baseline design, the authors trained parents in a group format and conducted observations of child compliance in the home throughout the intervention., Results: Observations indicated improvements in generalized child compliance that were maintained up to 6 months posttreatment. Pre-/post maternal reports indicated significant reductions in maternal perception of child noncompliance, externalizing and internalizing problems, and parenting stress., Conclusions: Researchers have noted a dearth of empirically supported interventions for children from violent homes. Errorless compliance training may help to fill this void, as it appears well suited to treatment of difficulties encountered by these youngsters and their parents.

The effects of the delta agonists SNC80 and deltorphin II on ambulation and rearing activity were measured in habituated and non-habituated rats. SNC80 (30, 100, 200, 400 nmol, i.c.v.) and deltorphin II (3, 15, 30, 60 nmol, i.c.v.) induced similar, dose-dependent biphasic locomotor effects in non-habituated subjects. An initial decrease in exploratory activity was associated with anxiogenic signs such as pilo-erection, freezing behaviour and pupil dilation for each drug. Pre-treatment with the delta antagonist naltrindole (10 nmol, i.c.v.) inhibited the depressant effect, but not the subsequent stimulant effect, on locomotor activity in response to 30 nmol deltorphin II in this assay (P<0.05). In habituated rats, deltorphin II (0.03, 0.1, 0.3, 3 nmol, i.c.v.) caused significant, naltrindole-reversible increases in locomotor activity (P<0.05 for all doses) at 1,000-fold lower doses than those required for a similar response to SNC80 (10, 30, 100, 300 nmol, i.c.v.). Pharmacokinetic studies suggest that these compounds penetrate the brain to similar extents following i.c.v. injection. The substantial potency difference between deltorphin II and SNC80 in stimulating locomotor activity in habituated rats suggests pharmacological heterogeneity for these delta opioid receptor agonists.

Objective: To assess executive function in girls with attention-deficit/hyperactivity disorder (ADHD) using oculomotor tasks as possible trait markers for neurobiological studies., Method: Thirty-two girls aged 6 to 13 years with DSM-IV ADHD and 20 age-matched, normal control girls were tested on a variety of oculomotor tasks requiring attention, working memory, and response inhibition, which included smooth pursuit, delayed response, and go-no go tasks., Results: Girls with ADHD performed the delayed response task correctly on 32% of trials as measured by number of memory-guided saccades, in contrast to 62% of trials for control subjects (p = .0009). Patients made twice as many commission errors to no go stimuli (p = .0001) and 3 times as many intrusion errors (saccades in the absence of go or no go stimuli; p = .004) during the go-no go task compared with controls. Smooth pursuit performance was statistically equivalent across subject groups. Repeated testing in a subgroup of 15 patients revealed substantial practice effects on go-no go performance., Conclusions: The data confirm that girls with ADHD exhibit impairments in executive function, as has been reported in boys, implying a similar pathophysiology of ADHD in both sexes. However, practice effects may limit the utility of the oculomotor go-no go task for some neurobiological studies.

Procedural sedation and analgesia are core skills in emergency medicine. Various specialty societies have developed guidelines for procedural sedation, each reflecting the perspective of the specialty group. Emergency practitioners are most likely to embrace guidelines developed by people who understand emergency department (ED) skills, procedures, conditions, and case mix. Recognizing this, the Canadian Association of Emergency Physicians (CAEP) determined the need to establish guidelines for procedural sedation in the ED. In March, 1996, a national emergency medicine (EM) working committee, representing adult and pediatric emergency physicians, was established. This committee teleconferenced with representatives of the Canadian Anesthetic Society (CAS) to identify problems, perspectives, and controversial issues, and to define a process for guideline development. The EM committee subsequently reviewed existing literature, determined levels of evidence, and developed the document, which evolved based on feedback from the CAS and CAEP Standards Committees. The final version was approved by the CAEP Standards Committee and the CAEP Board of Directors, then submitted for peer review. These guidelines discuss the goals, definitions, and principles of ED sedation, and make recommendations for pre-sedation preparation, patient fasting, physician skills, equipment and monitoring requirements, and post-sedation care. The guidelines are aimed at non-anesthesiologists practicing part-time or full-time emergency medicine. They are applicable to ED patients receiving parenteral analgesia or sedation for painful or anxiety-provoking procedures. They are intended to increase the safety of procedural sedation in the ED.

The aim of this article is to provide an evidence-based guideline on the management of acute migraine headaches in the Emergency Department setting. After a Medline search that covered 1965 to the present, all randomized controlled trials were reviewed. Recommendations as to the efficacy of abortive anti-migraine medications were based on the Canadian Medical Association's Guideline for Guidelines. Classes of medications that are discussed include: dopamine antagonists, serotonin agonists, opioids, local anesthetics, non-steroidal anti-inflammatory agents, and steroids. The recommendations are limited to discussing the efficacy of specific medications, adverse effects to be expected, as well as associated headache rates after discharge. Specific recommendations as to which medication might offer a superior treatment have not been proposed due to lack of proper comparative trials as well as lack of information on headache and quality of life after discharge from the Emergency Department.

A sensitive and selective method was developed for the simultaneous determination of chloroquine (CQ) and its desethylated metabolites monodesethylchloroquine (DCQ) and bisdesethylchloroquine (BDCQ) in human liver microsomes. Analytes were separated on a C1 column using methanol-water (70:30, v/v) and triethylamine (0.1% v/v) as the mobile phase. The fluorescence detector was set at 250 (excitation) and 380 nm (emission). Following protein precipitation with ice-cold acetonitrile, microsomal incubation supernatants were directly injected into the HPLC system. Typically, 200 microl of incubate were diluted with 200 microl of acetonitrile and 15 microl were injected. The limit of quantitation was 78 nM of CQ or metabolite. Intra-day variability averaged 2.9% for CQ, 1.5% for DCQ and 2.5% for BDCQ. Inter-day variability was 3.1% for CQ, 3.5% for DCQ and 3.7% for BDCQ. Mean accuracies were 100% for CQ and BDCQ and 102% for DCQ.

This review article focuses on the specificities of chiral liquid chromatography, with particular emphasis on stability, stereoconversion, enantiomeric separation, recovery and drug concentration determinations. In addition, the paper presents an overview of the different steps which have to be followed for a chiral method to be validated. Sensitivity, selectivity, linearity, precision and accuracy all have to be ensured for three chemical entities, the two enantiomers and the racemate. Only accurate and precise concentrations of the parent drug and its metabolites will lead to the reliable description of their in vitro stability and in vivo body disposition.

Dextromethorphan (DM) is O-demethylated into dextrorphan (DEX) in humans by the cytochrome P450 designated as CYP2D6 and N-demethylated into 3-methoxymorphinan (3MM) via CYP3As. Clinically, DM has been successfully used as an index of CYP2D6 and this paper describes analytical and clinical data that will help evaluate the use of DM hydrobromide as a probe of CYP3A activity. DM and its three demethylated metabolites were measured in a 4-h spot urine sample using a HPLC method employing solid-phase extraction (C(18)), analysis on a phenyl column [mobile phase, methanol-acetonitrile-phosphate buffer (10 mM, pH 3.5, 20:25:55, v/v)] and fluorescence detection (excitation at lambda=228 nm, no emission cut-off filter). The urinary molar ratio DM-DEX was used to assess CYP2D6 activity while DM-3MM was used for CYP3As. The DM-3MM ratios were sensitive to the co-administration of selective CYP3A inhibitors grapefruit juice and erythromycin. In addition, in healthy volunteers and cancer patients, the N-demethylation of DM correlated with the CYP3A-mediated metabolism of verapamil and tamoxifen. DM appears to be a promising way to simultaneously phenotype patients for CYP2D6 and CYP3As.

A prospective blinded observational study was carried out to document acute pain assessment and management in an academic emergency department. Over a 2-month period, 42 patients with a primary complaint of pain were studied. Physicians and nurses did not document levels of pain or changes in pain during patient stays. Eleven of 42 patients had severe pain upon arrival, 5 of whom received medications, only 1 obtaining good relief. No other patient received medication, although five others received some intervention for their pain. Eleven of 38 patients who were discharged home had severe pain on discharge. Despite minimal pain relief, patient satisfaction with pain management was relatively high. Pain assessment and treatment may be poorer than previous retrospective studies have indicated.

Pain is the most common presenting complaint heard in Emergency Medicine, yet it is poorly controlled. Evaluation of this pain should be with use of objective pain scales completed by the patient, not relying on physician impression. Treatment modalities available in the Emergency Department, a review of medications and their dosing as well as specifics to pediatric pain management are presented. The final section reviews situation or diagnosis specific pain control: headaches, renal colic, polytrauma victims, abdominal pain, soft tissue injury and acute arthritis. These recommendations are based on a Canadian Association of Emergency Physicians (CAEP) consensus conference held in April 1993. The literature was reviewed extensively and used as the basis for the consensus workshops and discussion. At the writing of the consensus paper, however, no specific ideas were borrowed from any one article. The appended bibliography is suggested reading, selected from the larger literature review. There are to date few controlled multi centre trials in overall pain management that would allow guidelines to be produced.

A high-performance liquid chromatographic assay coupled with electrochemical detection has been developed for the determination of vecuronium and its three putative deacetylated metabolites in human plasma. A novel solid-phase extraction procedure allowed good recovery of both vecuronium and its metabolites, together with ease and speed of execution. This method was sensitive, reproducible and accurate over the therapeutic range of concentrations of vecuronium and its metabolites, and was applied successfully to a study of the pharmacokinetics of vecuronium in anaesthetized patients.

Suxamethonium increases neuromuscular block produced by non-depolarizing agents administered subsequently. To determine if this effect has a pharmacokinetic or pharmacodynamic origin, 18 ASA physical status I or II adults received atracurium 0.2 mg kg-1, with (n = 10) or without (n = 8) previous injection of suxamethonium 1 mg kg-1, during a thiopentone-nitrous oxide-isoflurane (0.5% end-tidal) anaesthetic. Arterial blood samples were obtained and plasma atracurium concentration measured by HPLC. Train-of-four stimulation was applied to the ulnar nerve and the force of contraction of the adductor pollicis muscle was recorded. Mean (SEM) volume of distribution was slightly greater with previous suxamethonium (143 (13) ml kg-1) than without (109 (5) ml kg-1) (P less than 0.04). Mean elimination half-life was unaffected (20.3 (0.8) min and 20.4 (1.6) min, respectively). Neuromuscular block was more intense and recovery was slower with previous administration of suxamethonium. Atracurium concentration at 50% block (Cpss50) was 305 (30) ng ml-1 with and 454 (25) ng ml-1 without previous suxamethonium (P less than 0.01). It is concluded that suxamethonium may be associated with a slight increase in the volume of distribution of atracurium, but this effect is more than compensated by a decrease in atracurium concentration required for a given effect.

A high-performance liquid chromatographic method coupled with fluorometric detection has been developed for the determination of atracurium and its major end-product laudanosine in human plasma. The method enables good separation of atracurium from its metabolites after direct precipitation of plasma proteins. The assay is sensitive, reproducible and linear for atracurium concentrations ranging from 31.25 to 8000 ng/ml. In a clinical setting, drugs commonly administered during anesthesia did not interfere with the assay. This method provides a simple and time-saving alternative to existing methods.

The present study was undertaken to evaluate the state of glucocorticoid cytosol receptor binding in muscle hypertrophied by compensatory overload in normal, testosterone-treated, and castrated male rats. Compensatory hypertrophy in plantaris muscles was induced by myomectomy of the gastrocnemius and the soleus of 100 male Sprague-Dawley rats. The controlateral sham-operated limb was used as control. The rats were divided in 3 groups: control injected with peanut oil, castrated injected with peanut oil, and castrated injected with testosterone propionate (1 mg/rat). Five days later the animals were sacrificed and the muscles obtained for glucocorticoid receptor binding studies using [3H]dexamethasone as ligand. Receptor concentrations (Bmax) were significantly increased in hypertrophied muscle and the increase was not affected either by castration or by androgen treatment. These results tend to suggest that gonadal steroids do not participate in hypertrophy-induced proliferation of glucocorticoid receptors.