Viva la VOSCE?

BACKGROUND: The COVID-19 pandemic lockdown precluded face-to-face final Objective Structured Clinical Examinations (OSCE) in the UK. RESULTS: In response, we rapidly developed and then successfully implemented a novel Virtual Objective Structured Clinical Examination (VOSCE). CONCLUSIONS: In this article we both describe and reflect on our experience as well as discuss the implications for future undergraduate assessment as the situation evolves.

Temporal and spatial changes in cell wall composition in developing grains of wheat cv. Hereward.

A combination of enzyme mapping, FT-IR microscopy and NMR spectroscopy was used to study temporal and spatial aspects of endosperm cell wall synthesis and deposition in developing grain of bread wheat cv. Hereward. This confirmed previous reports that changes in the proportions of the two major groups of cell wall polysaccharides occur, with beta-glucan accumulating earlier in development than arabinoxylan. Changes in the structure of the arabinoxylan occurred, with decreased proportions of disubstituted xylose residues and increased proportions of monosubstituted xylose residues. These are likely to result, at least in part, from arabinoxylan restructuring catalysed by enzymes such as arabinoxylan arabinofurano hydrolase and lead to changes in cell wall mechanical properties which may be required to withstand stresses during grain maturation and desiccation.

Fruit juice authentication by 1H NMR spectroscopy in combination with different chemometrics tools.

To discriminate orange juice from grapefruit juice in a context of fraud prevention, (1)H NMR data were submitted to different treatments to extract informative variables which were then analysed using multivariate techniques. Averaging contiguous data points of the spectrum followed by logarithmic transformation improved the results of the data analysis. Moreover, supervised variable selection methods gave better rates of classification of the juices into the correct groups. Last, independent-component analysis gave better classification results than principal-component analysis. Hence, ICA may be an efficient chemometric tool to detect differences in the (1)H NMR spectra of similar samples, and so may be useful for authentication of foods.

Phase I trial of temozolomide using an extended continuous oral schedule.

Temozolomide, a methylating imidazotetrazinone, has antitumor activity against gliomas, malignant melanoma, and mycosis fungoides and is presently administered as a 5-day oral schedule every 4 weeks. This Phase I study aimed to determine the maximum tolerated dose of temozolomide administered as a single oral daily dose for a continuous 6- or 7-week period, evaluate the plasma pharmacokinetics on this schedule, and compare total plasma exposure over 7 weeks with the conventional 5-day regimen. Twenty-four patients with varying tumor types (17 of 24 gliomas) received temozolomide. All had clinically evaluable, refractory disease; normal renal, hepatic, and bone marrow function; and WHO performance status < or = 2. Temozolomide was administered at 50 mg/m2/day, increasing by 25 mg/m2/day/cohort until at 100 mg/m2/day grade 4 myelotoxicity forced dose reductions to 85 mg/m2/day, then 75 mg/m2/day. At 75 mg/m2/day the regimen was extended to 7 weeks, allowing the future potential combination with irradiation for primary gliomas. Patient responses (standard Union International Contre Cancer criteria; for gliomas objective response) and toxicity were assessed. Temozolomide plasma pharmacokinetics were determined on day 1 and at the beginning of the final week of administration (n = 5). The most frequent toxicities were myelosuppression and grades 1 and 2 nausea and vomiting. Grade 4 leucopenia and thrombocytopenia occurred in one of four patients receiving 100 mg/m2/day temozolomide and in one of seven patients receiving 85 mg/m2/day. These hematological toxicities did not exceed grade 2 in 10 patients receiving 75 mg/m2/day temozolomide. One of 4 malignant melanoma patients and 7 of 17 glioma patients (41%) demonstrated tumor responses. The overall response rate for this prolonged schedule was 33% (objective response, 7 of 24 patients; partial response, 1 of 24 patients); also, 6 of 17 glioma patients maintained SD. Peak plasma temozolomide concentrations were obtained 30-90 min after oral administration. Elimination in plasma was best described by a monoexponential equation with an elimination half-life of 96 +/- 16 min. No plasma accumulation of temozolomide occurred. Toxicity was greatest in higher dose cohorts, with a resultant maximum tolerated dose of 85 mg/m2/day, whereas lower dose cohorts tolerated the schedule well. The area under the temozolomide plasma versus time curve was noncumulative between the first and last week of the schedule. Temozolomide administration of 75 mg/m2/day over a 7-week period permits a 2.1-fold greater drug exposure/4 weeks in comparison with the 5-day schedule of 200 mg/m2/day repeated every 28 days. The overall response rate was 33% (glioma patients, 41% and a further 25% SD). Temozolomide (75 mg/m2/day) for 7 weeks is the recommended starting dose for further assessment of this schedule.

Authentication of beef versus horse meat using 60 MHz 1H NMR spectroscopy.

This work reports a candidate screening protocol to distinguish beef from horse meat based upon comparison of triglyceride signatures obtained by 60 MHz (1)H NMR spectroscopy. Using a simple chloroform-based extraction, we obtained classic low-field triglyceride spectra from typically a 10 min acquisition time. Peak integration was sufficient to differentiate samples of fresh beef (76 extractions) and horse (62 extractions) using Naïve Bayes classification. Principal component analysis gave a two-dimensional "authentic" beef region (p=0.001) against which further spectra could be compared. This model was challenged using a subset of 23 freeze-thawed training samples. The outcomes indicated that storing samples by freezing does not adversely affect the analysis. Of a further collection of extractions from previously unseen samples, 90/91 beef spectra were classified as authentic, and 16/16 horse spectra as non-authentic. We conclude that 60 MHz (1)H NMR represents a feasible high-throughput approach for screening raw meat.

Temozolomide: a new oral cytotoxic chemotherapeutic agent with promising activity against primary brain tumours.

Temozolomide, a new oral cytotoxic agent, has been given to 28 patients with primary brain tumours. Treatment was given at a dose of 150 mg/m2/day for 5 days (i.e. total dose 750 mg/m2) escalating, if no significant myelosuppression was noted on day 22, to 200 mg/m2/day for 5 days (i.e. total dose 1000 mg/m2) for subsequent courses at 4 week intervals. A major improvement in computer tomography (CT) scan was noted in 5/10 patients with astrocytomas recurrent after radiotherapy, with a major clinical improvement but minor improvement on CT scan in one further patient. Reduction in the size of the CT lesion was also observed in 4/7 patients with newly diagnosed high grade astrocytomas given 2-3 courses of temozolomide prior to irradiation. 1 patient with recurrent medulloblastoma had a clinical response in bone metastases. Temozolomide was well tolerated with little subjective toxicity and usually predictable myelosuppression and is a promising new drug in the treatment of primary brain tumours.

The Charing Cross Hospital experience with temozolomide in patients with gliomas.

Temozolomide, a new oral cytotoxic agent, was given to 75 patients with malignant gliomas. The schedule used was for the first course 150 mg/m2 per day for 5 days (i.e. total dose 750 mg/m2), escalating, if no significant myelosuppression was noted on day 22, to 200 mg/m2 per day for 5 days (i.e. total dose 1000 mg/m2) for subsequent courses at 4-week intervals. There were 27 patients with primary disease treated with two courses of temozolomide prior to their radiotherapy and 8 (30%) fulfilled the criteria for an objective response. There were 48 patients whose disease recurred after their initial surgery and radiotherapy and 12 (25%) fulfilled the criteria for an objective response. This gave an overall objective response rate of 20 (27%) out of 75 patients. Temozolomide was generally well tolerated, with little subjective toxicity and predictable myelosuppression. However, the responses induced with this schedule were of short duration and had relatively little impact on overall survival. In conclusion, temozolomide given in this schedule has activity against high grade glioma. However, studies evaluating chemotherapy in primary brain tumours should include a quality-of-life/performance status evaluation in addition to CT or MRI scanning assessment.

Lung preservation: a review of current practice and future directions.

During the past 10 years, pulmonary transplantation has emerged as a successful mode of surgical therapy for suitable patients with end-stage lung disease. Current preservation techniques of donor lungs for subsequent transplantation include core-cooling and single flush perfusion. The relative merits of these are described. These methods are essentially restricted to 6 hours of ischemia. Research in lung preservation is aimed not only at extending the safe period of ischemia but also at improving the quality of preservation. Areas of interest include the ideal composition of the perfusate, relevant pharmacologic additives, and the best conditions for preservation and harvesting. Advantages and disadvantages of the various animal models are listed in addition to the methods used in assessing the quality of preservation. There have been major advances in experimental lung preservation during the past 10 years, and we are possibly on the threshold of incorporating some of these into clinical practice. Among the most important are the adoption of colloid-based perfusates, the more widespread use of free radical scavengers, and the use of leukocyte depletion.

Airway complications after pulmonary transplantation.

Airway healing was identified initially as one of the fundamental limitations of pulmonary transplantation. Recent experience suggests that this is no longer the case. A series of 67 pulmonary transplants (27 heart-lung, 31 single-lung, 9 double-lung) in 66 patients surviving more than 14 days was reviewed with reference to airway complications. There were 75 anastomoses at risk in two groups as defined by anastomotic location: 47 anastomoses in 38 patients in a bronchial group and 28 anastomoses in 28 patients in a tracheal group. A total of 10 airway complications developed (stenosis in 5 patients [4 bronchial group, 1 tracheal group] and dehiscence in 5 patients [1 bronchial group, 4 tracheal group]) causing two airway-related deaths (2 of 67) in the series. However, no significant correlation could be identified with either ischemic interval, suture technique, type of wrap, preoperative or postoperative steroid therapy, or date of first rejection episode. Airway complications are no longer a major limitation of pulmonary transplantation. Satisfactory airway healing can occur in both the presence of steroid therapy and the absence of an omental or pericardial wrap.

Multicentre CRC phase II trial of temozolomide in recurrent or progressive high-grade glioma.

PURPOSE: Patients with progressive or recurrent supratentorial high-grade gliomas were entered into a multicentre phase II trial to evaluate the efficacy and toxicity of temozolomide. METHODS: The treatment schedule was 150-200 mg/m2 per day orally for 5 days repeated every 28 days. Response evaluation was by a combination of neurological status evaluation (MRC scale) and imaging. RESULTS: Of 103 eligible patients enrolled, 11 (11%) achieved an objective response and a further 48 (47%) had stable disease. The median response duration was 4.6 months. Response rates were similar for anaplastic astrocytomas (grade III) and glioblastoma multiforme (grade IV) tumours. Predictable myelosuppression was the major toxicity. CONCLUSIONS: The observation of objective responses and tolerable side effects in this heterogeneous population of patients supports the further investigation of this agent in high-grade gliomas.

Distal tracheal stenosis in two children with double aortic arch and associated tracheomalacia.

Two case reports are presented in children who developed significant respiratory distress due to persistent tracheomalacia despite vascular decompression for double aortic arch. Cardiopulmonary bypass is suggested as a useful approach to the distal tracheal segment in children.

Arrhythmia prophylaxis after coronary artery surgery. A randomised controlled trial of intravenous magnesium chloride.

One hundred and thirty patients undergoing elective myocardial revascularisation were randomised to receive an intravenous infusion containing either 50 mmol magnesium chloride or placebo during the first 48 h following surgery. Magnesium was given to 66 patients and a placebo infusion to 64 patients. Postoperative serum magnesium concentrations fell in the placebo group (from 0.83 +/- 0.06 to 0.79 +/- 0.11 mmol/l) but were elevated in the magnesium group (from 0.82 +/- 0.05 to 1.2 +/- 0.25 mmol/l). In total, 35 patients (26.9%) had a supraventricular arrhythmia, of which 11 were in the magnesium group (16.7%) and 24 (37.5%) in the placebo group (P = 0.013). Three patients in the magnesium group had an episode of ventricular fibrillation and three patients in the placebo group had frequent ventricular ectopics. There was one death in the magnesium group associated with a perioperative myocardial infarction. This study shows that intravenous magnesium supplements reduce the incidence of supraventricular arrhythmias following coronary artery surgery.

An analysis of referrals for primary diagnostic abdominal ultrasound to a general X-ray department.

Abnormalities were found in 30% of examinations. Comparison of outpatient and family practitioner referrals in cases of suspected gallstones (24%:27%) indicated the value of GP access. There was also evidence, in the absence of a dilated common bile duct, that there was little value in extending the examination of the gallbladder. In post-cholecystectomy syndrome, endoscopic retrograde cholangiography is shown to be the investigation of choice. Abnormalities revealed in lower abdomen examinations were related only to pelvic organs and in this group examination should therefore be confined to this area. Pick-up rates of 15% were found in patients with left upper quadrant pain and a number of miscellaneous conditions. There is a marked increase in positive findings in the presence of a palpable mass.

The assessment of carotid and vertebral arteries: a comparison of CFM duplex ultrasound with intravenous digital subtraction angiography.

The use of duplex ultrasound with colour flow mapping is compared with that of intravenous digital subtraction angiography (IVDSA) in the investigation of carotid and vertebral artery disease. Ninety-nine carotid and vertebral arteries were examined independently by IVDSA and ultrasound for location and degree of disease in 30 men (mean age 58) and 20 women (mean age 53). For purposes of comparison the carotid tree was divided into six segments. In 46% of cases no disease was detected by either modality. In 36% of cases where disease was found, ultrasound found mild disease in arteries reported as normal by IVDSA. Both modalities detected the six cases of total occlusion found in the study. IVDSA underestimated five cases of major disease found by ultrasound at the bifurcation or in the bulb. Overall agreement in grading vessel segments was good, with 74.5% in complete agreement and 90.3% grading stenoses to within +/- 25% of the other modality. There was good qualitative agreement in the findings for vertebral arteries. In four patients conventional arteriography was also available for comparison.

NMR studies of acetan and the related bacterial polysaccharide, CR1/4, produced by a mutant strain of Acetobacter xylinum.

Acetan is a bacterial polysaccharide produced by Acetobacter xylinum NRRL B42. Chemical mutagenesis of A.xylinum allowed selection of a mutant strain which produced a new polysaccharide, CR1/4. 2D NMR methods have been used to assign the 1H and 13C spectra of the two polysaccharides and to determine that CR1/4 has the structure shown below. The total number of O-acetyl groups is slightly less than two per repeating unit. [formula: see text] The pentasaccharide side chain of acetan is truncated to a disaccharide unit in CR1/4, but the structures are otherwise identical. In particular, the degree of acetylation is about the same and the O-acetyl groups are located at the same position in both polysaccharides.

Isolation and characterization of rhamnogalacturonan oligomers, liberated during degradation of pectic hairy regions by rhamnogalacturonase.

Digests of modified hairy regions of apple pectin (MHR) obtained after degradation by rhamnogalacturonase (RGase) were analyzed for oligomer composition using high-performance anion-exchange chromatography and pulsed amperometric detection. A series of oligomers which appear to be characteristic of RGase degradation could be recognized. These oligomers were isolated on a preparative scale by size-exclusion chromatography and preparative anion-exchange chromatography and analyzed for sugar composition. 1H NMR spectroscopy showed that the oligomers consisted of between 4 and 9 sugar units with a backbone of alternating rhamnose and galacturonic acid residues, partly substituted with galactose residues linked to C-4 of the rhamnose moiety. The HPLC elution pattern showed that higher oligomers were also formed during incubation with RGase. These have the same basic structure but may contain other sugar units in addition to those given above. The oligomer composition of RGase digests of MHR isolated from apple, pear, leek, onion, carrot, and potato was very similar. Using anion-exchange chromatography to monitor the degradation of MHR at increasing incubation times, it was found that all the oligomers were present from the initial stages of the enzyme reaction and that the ratio between the different oligomers remained constant with time. Implications of these results for the structure of MHR and the mechanism of RGase action are discussed.

Structure identification of feruloylated oligosaccharides from sugar-beet pulp by NMR spectroscopy.

1D NMR (1H and 13C) and 2D NMR spectroscopy have been used to determine the structure of feruloylated oligosaccharides obtained by enzymic degradation or mild acid hydrolysis of sugar-beet pulp. Feruloylated oligosaccharides derived from pectic neutral side-chains containing arabinose or galactose residues were identified. In the feruloylated arabinose oligosaccharides, feruloyl groups were linked to O-2 of L-Ara f residues. The structure of the feruloylated arabinose disaccharide was identified as O-[2-O-(transferuloyl)-alpha-L-Ara f]-(1-->5)-L-Ara f and that of the feruloylated arabinose trisaccharide as O-alpha-L-Ara f-(1-->3)-O-[2-O-(trans-feruloyl)-alpha-L-Ara f]-(1-->5)-L- Ara f. The structure of the feruloylated galactose disaccharide was identified as O-[6-O-(trans-feruloyl) -beta-D-Gal p]-(1-->4)-D-Gal p. From our results, we suggest that the feruloyl groups present in sugar-beet pulp are linked to the arabinofuranosyl residues of the main core of alpha-(1-->5)-linked arabinan chains and to the galactopyranosyl residues of the main core of beta-(1-->4)-linked type I galactan chains.

Sucrose octabenzoate: assignment of 13C and 1H resonances of the sucrose moiety and the 13C resonances of the carbonyl carbons. Use of 13C-n.m.r. spectroscopy for the study of selective deacylation.

Assignment of the 1H and 13C signals arising from the carbohydrate portion of sucrose octabenzoate has been achieved using homonuclear shift correlation experiments (COSY) and one-bond 1H-13C heteronuclear shift correlation measurements, respectively. The 13C resonances of the carbonyl carbon atoms of the eight benzoyl groups are readily distinguished for solutions in benzene-d6-pyridine-d5 (1:1), and have been assigned by means of three-bond 1H-13C shift correlation studies coupled with measurement of the 13C-n.m.r. spectrum of a sucrose octabenzoate specifically labelled with 13C in some of the carbonyl groups. With this assignment, products of partial deacylation of the octabenzoate may readily be identified by treatment with excess of benzoyl-carbonyl-13C chloride followed by measurement of the 13C-n.m.r. spectrum of the labelled sucrose octabenzoate, so prepared, in the carbonyl region.

Identification by n.m.r. spectroscopy of oligosaccharides obtained by treatment of the hairy regions of apple pectin with rhamnogalacturonase.

2D-N.m.r. methods have been used to determine the composition of a mixture of oligosaccharides obtained by enzymic degradation of the modified hairy (ramified) regions of apple pectin with a new rhamnogalacturonase. The structures of the oligosaccharides were based on the unit alpha-Rhap-(1----4)-alpha-GalA-(1----2)-alpha-Rhap-(1----4)- GalA. A-beta-Galp unit was 4-linked to approximately half of the terminal Rhap residues and to half of the (1----2)-linked Rhap residues. The sample contained a mixture of a tetrasaccharide, two pentasaccharides, and one hexasaccharide.

Structure and conformation of a novel genetically engineered polysaccharide P2.

A new exocellular polysaccharide (P2) has been produced by the manipulation of a glycosyl transferase gene (aceP) involved in the biosynthesis of the polysaccharide acetan by the bacterium Acetobacter xylinum strain CKE5. The P2 polysaccharide has been studied by methylation analysis, reductive cleavage, and 1H and 13C NMR spectroscopy. The data are consistent with the structure predicted when the aceP gene is deactivated: [Molecular structure: see text]. The effect of cooling on proton NMR line width indicates a coil-helix transition in P2 at about 70 degrees C.