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This paper describes a set of learning outcomes that clearly define the abilities of medical graduates from any of the five Scottish medical schools. The outcomes are divided into 12 domains that fit into one of three essential elements for the competent and reflective medical practitioner.

The influence of antiepileptic drug monitoring on site at the epilepsy clinic was assessed in a prospective way by recording physicians' decisions before and after the drug concentration became available over the five-year period including 1986 to 1990. A total of 2,857 assays (1,121 carbamazepine, 798 phenytoin, 622 sodium valproate, and 72 phenobarbital) were performed during 2,696 hospital attendances on 618 epileptic patients. A gradual reduction in the proportion of visits monitored annually was observed (1986: 42%; 1987: 51%; 1988: 44%; 1989: 39%; 1990: 28%; p < 0.01). In 481 (17.8%) visits a change in management resulted from knowledge of the drug concentration. This was not an automatic response, as a large minority of unexpected results did not produce such an alteration. The proportion of decisions to adjust the drug dose after an unexpected result, however, tended to increase over the years of the study (1986: 43%; 1987: 70%; 1988: 60%; 1989: 78%; 1990: 54%; p < 0.01). Changes in anticonvulsant dose following an unexpected result were observed in "low" (59%), "therapeutic" (61%), and "high" (52%) concentration categories. Possible benefit for patients can be inferred from the observation that more assay results fell within the target range in the last (61%) compared with the first (33%) year of monitoring (p < 0.01).

To assess the value of on site therapeutic drug monitoring at the epilepsy clinic, management decisions were recorded before and immediately after antiepileptic drug (AED) concentrations became available. In the first year of this prospective study, 632 [277 carbamazepine (CBZ), 170 phenytoin (PHT), 113 valproate (VPA), and 72 phenobarbital (PB)] assays were performed during 488 clinic attendances in 182 actively managed epileptic patients. The results of drug analysis led to alterations in management at 114 patients visits, i.e., 23% of those monitored. Dosage was increased in response to the circulating AED concentration in 12% of consultations and decreased in another 7.5%. Unsuspected poor compliance was uncovered in eight patients, and in three others an AED was added or discontinued on the basis of the assay result. The time of the next appointment was rearranged in 58 attendances. Only 50% of results were in the "therapeutic" ranges for the four major AEDs. Dosage was adjusted (50 up, 16 down) after 54% of low results. "Therapeutic" levels were followed by a change in AED dose (52 up, 31 down) in 26%. Only 29% of concentrations above the "therapeutic" range persuaded the doctor to alter the dosage regimen, and in 20% of these an increase in dose was recommended. On-site AED monitoring had an immediate impact on clinical decision-making in greater than 23% of consultations but in a form more subtle than the simple quest for a therapeutic result.

Our objective was to determine whether the fluorescence of skin collagen, which may reflect the accumulation of advanced glycosylation end products, is increased in young patients with type I (insulin-dependent) diabetes. Our study design was a cross-sectional case-control study in a referral-based diabetic clinic in an academic hospital. Study subjects comprised a convenience sample of 18 type I diabetic patients aged 17-30 yr and 8 age-matched healthy control subjects. The fluorescence of collagen was measured in skin biopsy material. Collagen-linked fluorescence (CLF) was increased in diabetic patients (mean 10.5 [range 5.8-15.8] U/mg) compared with control subjects (7.6 [5.6-10.1] U/mg, P less than 0.02). In diabetic patients, CLF was related to age (r = 0.581) and duration of diabetes (r = 0.697) but not concentration of glycosylated hemoglobin (r = 0.082). Partial correlation analysis demonstrated that duration of diabetes is the main factor determining the fluorescence of collagen in these patients. There was a relationship between CLF and presence of diabetic retinopathy after the data were adjusted for patient age and duration of diabetes (P = 0.023). Increased fluorescence of skin collagen can be detected in young type I diabetic patients and is primarily related to duration of diabetes.

Serum bile salt measurements and intravenous clearance of glycocholate were performed in a woman with Dubin-Johnson syndrome. Fasting conjugated cholate concentration was raised and prolonged intravenous clearance of sodium glycocholate revealed a secondary rise in conjugated cholate concentration after two hours. The intravenous clearance of bromsulphthalein also showed a secondary rise. These findings support the proposal that Dubin-Johnson syndrome is not homogeneous and that in some patients a abnormality of bile salt clearance coexists with an abnormality of bilirubin nd bromsulphthalein clearance.

1. The subcellular distribution of conjugates of cholic acid and chenodeoxycholic acid between cytosol, nuclei, mitochondria and microsomes in rat liver has been determined. 2. The partition coefficients for the distribution of these bile acids between subcellular fractions and buffer have been measured and used to construct a compartmental model of the amounts of conjugated bile acids present in the different subcellular organelles in vivo. 3. This model indicates that a large percentage of the bile acid in the rat liver is found in the nuclear fraction; 42% of the cholic acid conjugates and 27% of the chenodeoxycholic acid conjugates. Substantial amounts of bile acid are also present in microsomes and mitochondria suggesting that published estimates of the amounts of bile acids in these fractions are underestimates. 4. The model also allows the amount of bile acid which is in free solution in cytosol to be determined; 10.9% of the cholic acid conjugates and 4.1% of the chenodeoxycholic acid conjugates in rat liver were present in this fraction. Knowlege of the amount of free bile acid allows possible roles of the cytosolic bile binding proteins to be assessed.

Duodenal bile salt concentrations were measured throughout one day in six patients with primary biliary cirrhosis while they were eating a normal ward diet. Five of them had lost weight; none had ascites. Each patient had a radiologically normal small bowel and a normal jejunal biopsy. No clear relationship between high faecal fat excretion and abnormally low duodenal bile salt concentration was found. Xylose absorption was abnormal in five patients. If weight loss in primary biliary cirrhosis is due to malabsorption, factors other than a reduced small intestinal bile salt concentration must be important.

1. The binding of cholic acid to 100000g supernatants from rat livers was investigated by equilibrium dialysis and gel-exculsion chromatography. 2. Supernatants were found to contain at least two classes of binding site for cholic acid. 3. These recptor molecules are probably proteins since incubation with proteolytic enzymes resulted in complete loss of cholic acid binding. 4. Supernatants were added to columns of Sephadex G-75, and two groups of fractions were shown to bind cholic acid. One of these contained low-affinity binding sites and the other contained both low- and high-affinity binding sites. 5. Feeding cholestyramine had no effect on cholic acid binding. 6. Increased cholic acid binding occurred after injection of phenobarbitone. There was an increase in the amount of the low-affinity component but no change in the high-affinity component. 7. The dissociation constants of the binding of cholic acid suggest that the binding proteins may be involved in bile acid transport.

The concentrations of conjugated cholate and chenodeoxycholate have been measured in samples of rat thoracic duct lymph, portal and systemic blood. Both these bile salts were present in lymph. After administration of a corn oil meal the flux of both these bile salts in lymph increased significantly (P less than 0.001; Student's t test), the cholate flux rising from 0.9 +/- 0.6 nmol/h (mean +/- SD) to 3.5 +/- 1.1 nmol/h postprandially and the chenodeoxycholate flux rising from 2.5 +/- 0.8 to 4.6 +/- 1.6 nmol/h. This postprandial increase in bile salt flux was due to both an increase in bile salt concentration and an increased lymph flow which rose significantly (P=0.007) from 0.5 +/- 0.2 to 0.7 +/- 0.2 ml/h. Biliary drainage significantly (P less than 0.001) reduced the flux of cholate and chenodeoxycholate in lymph to 0.08 +/- 0.05 and 0.17 +/- 0.10 nmol/h respectively. These biliary drained animals produced no significant rise (P greater than 0.1) in bile salt flux when fed the corn oil meal. The ratio of the concentrations of conjugated cholate to chenodeoxycholate in systemic and portal blood was 1.9:1 and 3.3:1 respectively. In contrast, this bile salt ratio was only 0.44:1 in lymph. These studies show that bile salts, in particular chenodeoxycholate, can pass directly from the intestine into lymphatics thus establishing an enterolymphatic circulation of bile salts.

Several reports have been published commending the use of 14C-labelled triglyceride breath tests in the assessment of fat malabsorption. We report further studies using gyceryl [14C]tripalmitate. Corrections for age, weight or metabolic rate failed to improve the test's ability to discriminate between malabsorbers and control subjects. A correction for respiratory quotient improved the linear correlation observed between the breath test results and daily faecal fat excretion. The significance of these findings is discussed and a number of problems identified which, at present, are preventing the introduction of breath tests for fat malabsorption into routine clinical practice.

Cholic acid-binding activity in cytosol from rat livers appears to be mainly associated with enzymes having glutathione S-transferase activity; at least four of the enzymes in this group can bind the bile acid. Examination of the subunit compositions of different glutathione S-transferases indicated that cholic acid binding and the ability to conjugate reduced glutathione with 1,2-dichloro-4-nitrobenzene may be ascribed to different subunits.

1. Rats were maintained in a strictly controlled environment of 12 h illumination and 12 h darkness. At regular intervals during the light/dark cycle the portal blood conjugated cholic acid and conjugated chenodeoxycholic acid concentrations were measured. The bile salt concentrations exhibited similar diurnal rhythms, the highest concentrations occurring in the middle of the dark phase. 2. The concentrations of conjugated cholic and chenodeoxycholic acids in the portal blood of rats fed a diet containing the bile salt sequestrant, cholestyramine, were significantly lower than those found in rats given a control diet. 3. During total biliary drainage the portal blood concentrations of conjugated cholic and chenodeoxycholic acids fell to a minimum 6--8 h after the start of the experiment, whereas bile salt synthesis in hepatocytes isolated from the rats was not increased until the least 13 h after the commencement of total biliary drainage. 4. These results suggest that the concentrations of bile salts in the portal blood do not affect directly the diurnal fluctuation in rates of bile salt synthesis, as the response of synthesis to a change in portal blood bile salt concentrations is too slow. 5. When the rats were given a small supplement of cholesterol in the diet to suppress hepatic cholesterologenesis prior to being subjected to total biliary drainage, the changes in the portal blood concentrations of conjugated cholic and chenodeoxycholic acids and the synthesis of the two bile salts by isolated hepatocytes were similar to those found in rats given the control diet. 6. The rise in bile salt production seen during biliary drainage may not be dependent exclusively on a preceding increase in cholesterol synthesis.

Plasma 5-S-cysteinyldopa (5-S-CD) concentration measured in healthy volunteers in Edinburgh, Scotland (latitude 56 degrees N) showed only minor changes during the day. However, when measurements were performed over a 12-month period a significant rise in 5-S-CD concentration was found. Skin pigmentation and hair colour were not related to plasma 5-S-CD levels. Patients with psoriasis treated with ultraviolet-B or photochemotherapy (PUVA) developed an almost two fold increase in their plasma 5-S-CD level within the first five treatments, before pigmentation developed, subsequent increments of up to four times the pretreatment level being found in the PUVA group. Dithranol treatment caused an increase in plasma 5-S-CD in some psoriatic patients, suggesting a possible association between skin erythema and elevated 5-S-CD levels. The value of plasma 5-S-CD in the follow-up of patients with malignant melanoma does not seem to be invalidated by unavoidable exposure of the subjects to sunlight in a temperate climate such as that of South East Scotland.

A general method for the preparation of 125I-labelled bile acid-histamine or 125I-labelled bile acid-tyramine conjugates is presented. The method is simple, quick and produces ligands in good yield (30%). The characteristics of a radioimmunoassay for conjugated chenodeoxycholic acid, based on an 125I-labelled ligand prepared by the method, are also described. The assay produced values for fasting serum concentrations of conjugated chenodeoxycholic acid that agree well with previous data.

As part of a detailed study into the solubility of calcium in bile, the contribution of the biliary lipids, bile salt, lecithin, and cholesterole to calcium binding in bile has been assessed. Micellar binding could account for 80% of the calcium binding in hepatic bile but only 50% in gallbladder bile. It is suggested that such binding, in soluble form, lowers the activity of calcium in bile, and therefore its liability to precipitation as gallstone nuclei.

A modification of the manual glucose oxidase-gum guaiacum method of Shipton, B., Wood, P.J. and Marks, V. ((1975) Med. Lab. Tech. 32,33), for use with plasma, is described. The method enables rapid measurement of plasma or serum glucose concentration between 1 and 22 mumol/1 using 10 mul of sample and a single colour reagent. Experience of the method in routine use has shown that the coefficient of variation, beteen batch, is 2.1% and the time taken for analysis of 1 sample, standards and a quality of control sample is 12 min (including centrifugation of blood to obtain plasma).

A 125I-labelled photolabile derivative of cholic acid has been used to investigate the organic anion-binding Y' fraction from rat liver, prepared by the method of Sugiyama, Y., Yamada, T. and Kaplowitz, N. (1982) Biochimica Biophysica Acta, 709, 342-352. The use of this photoaffinity probe led to the discovery of previously undescribed bile acid-binding proteins. A comprehensive purification scheme for the Y' proteins which allows the isolation of these novel binding species is described. Electrophoretic analysis shows that the Y' binders can be divided into two groups. The proteins in group 1 are dimeric and the 5B, 6E and 7F binding species consist of subunits with approximate molecular masses of 19.6, 15.6 and 14.9 kDa, respectively. The group 2 binding proteins, 5C, 5D and 8C, are monomeric and have molecular masses of approximately 36.2, 36.2 and 33 kDa, respectively. Calculation of the incorporation of 125I by these proteins showed that the group 1 proteins displayed a significantly greater specific incorporation of radioactivity than group 2. The specificity of 125I-labelled 3 beta-azidocholylhistamine is further demonstrated by analysis of tryptic digests of photoaffinity labelled Y' binders and glutathione S-transferases AA, A, D and F by reverse-phase high-performance liquid chromatography (HPLC) and thin-layer chromatography (TLC). The majority of the radioactivity was shown to be incorporated into a single component, which was not coincident with the free photoaffinity label.

The two dimeric lithocholic acid-binding proteins previously identified as ligandin (YaYa) and glutathione S-transferase B (YaYc) were isolated from rat liver cytosol. These proteins have molecular weights of 44000 and 47000 respectively. The recovery of these two proteins from liver was not affected by the addition of the proteinase inhibitor Trasylol. No spontaneous interconversion between these two proteins was observed on storage. YaYa and YaYc proteins yielded peptides of identical molecular weight after limited digestion with Staphylococcus aureus V8 proteinase. Analytical and preparative tryptic-digest peptide 'maps' showed that all the soluble peptides obtained from YaYa protein were also recovered from YaYc protein. Approximately six extra soluble peptides, which were not recovered from YaYa protein, were obtained from the tryptic digest of YaYc protein. Subdigests of the insoluble tryptic-digest 'cores' also resulted in the recovery of identical peptides from both proteins. Evidence is presented that the Ya subunit possessed by both proteins is identical; glutathione S transferase B is a hybrid of ligandin and glutathione S-transferase AA. The Ya monomer is responsible for lithocholate binding.

A constant-volume ultrafiltration technique is described, and details of its assessment presented. The retention characteristics of two membranes were evaluated using molecules of known molecular weight. The technique is rapid, precise, economical of material and yields equilibrium data. In these respects, it compares favourably with conventional techniques such as equilibrium dialysis.

Perioperative digoxin concentrations were measured in 20 unselected adult patients undergoing coronary surgery. None of the patients were receiving treatment with digoxin. A digoxin-like immunoreactive substance was found in 16 patients postoperatively. This substance, if pharmacologically active, may have important clinical implications in the management of patients after open heart surgery.

1. The effect of systematic error (loss of ligand, complex or macromolecule) on three of the experimental designs by which equilibrium dialysis may be used to quantify the interaction of ligand and macromolecule is examined theoretically, and the design that is least sensitive to systematic error is identified. 2. Thirteen methods for fitting the binding isotherm to experimental data are compared by using them to analyse simulated data containing random error, and the most reliable method is identified.

Liver function was assessed in 38 Edinburgh haemophiliacs. Results before the introduction of NHS intermediate factor VIII concentrate from 1974 onwards were compared with values in 1979. Measurements of serum bile salts in 16 patients as well as conventional liver function tests gave useful evidence of deranged liver function. Deterioration over the five-year follow-up period was seen only in patients on home treatment using large amounts of factor VIII concentrate, and there was no association between cryoprecipitate usage and derangement of liver function.

The binding of bile salts to proteins in rat plasma and rat lymph has been investigated. Under the non-equilibrium conditions of gel chromatography no binding of glycochenodeoxycholate or glycocholate to any of the lymph proteins was observed. In contrast, plasma bound a proportion of both bile salts. When lymph was treated with charcoal, binding of glycochenodeoxycholate to a protein with a molecular weight identical to albumin occurred. Equilibrium binding studies showed that the binding of glycocholate to partially purified plasma albumin exhibited saturation kinetics with a dissociation constant of 2 x 10(-4) M. In contrast, the binding of glycocholate to lymph albumin was non-saturable. Potassium oleate, when added to plasma in a free fatty acid : albumin molar ratio of 3.8 : 1, almost completely inhibited the binding of chenodeoxycholate to plasma albumin. The endogenous free fatty acid : albumin ratios found in systemic plasma and lymph were 0.6 : 1 and 9.2 : 1, respectively. It is suggested that the high free fatty acid concentrations found in lymph inhibit the binding of bile salts to albumin.

Fasting and postprandial serum bile salt concentrations and intravenous glycocholate disappearance were studied in 20 patients with anicteric liver disease who had only minor abnormalities of conventional liver function tests. Abnormalities in the fasting or two hour postprandial conjugated cholate concentrations were found in all but one of the patients who had an abnormality in bilirubin concentration. In these patients, fasting and postprandial conjugated cholate concentrations were raised on average three and two times respectively above the upper limit of the reference range, while bilirubin concentration was raised only 50%. Postprandial conjugated cholate concentrations were also abnormal in two patients with normal bilirubin concentrations. Measurement of fasting and postprandial conjugated chenodeoxycholate concentration and intravenous glycocholate disappearance proved less informative than the fasting and postprandial conjugated cholate test. These results suggest that, where bilirubin concentrations are normal or only slightly raised, measurement of serum fasting and two hour postprandial conjugated cholate concentrations may prove helpful in the detection of minor abnormalities in hepatic anion transport.

1. Normally fed and cholestyramine-treated rats were injected through the superior mesenteric vein with different amounts of radiolabelled glycoholic acid and the appearance of radioactivity in bile was measured. 2. In normally fed rats radioactivity appeared in bile within 30 s of injection and reached a maximum after 2 1/2 min; in the cholestyramine-treated animals the appearance of radioactivity was slower and less of the injected material was excreted into bile. 3. At 10 min after injection, livers were removed and the amounts of radioactive glycoholic acid and endogenous cholic acid conjugates in nuclei and cytosol were determined; most of the bile acid was found in the cytosol, only small amounts being found in nuclei. 4. Nuclear preparations from both normally fed and cholestyramine-fed rats were extracted with KCl (0.4 M) in an attempt to identify a putative bile acid receptor, but no such receptor was found. 5. Regulation of bile acid synthesis does not involve nuclear binding of bile acids.

A frame-based computer-aided learning system for use on microcomputers has been developed for teaching clinical chemistry. The system presents a case-study model that can be used by students to assess their ability to make a diagnosis, to select patterns of investigations and to institute treatment.

We have studied serum fasting and postprandial primary bile salt concentrations in a group of 10 consecutive hyperlipidaemic subjects. The efficiency of hepatic bile salt clearance in the same subjects was aslo studied using an injected dose of sodium glycocholate. No increase in serum fasting or postprandial concentrations of the primary bile salts were observed and hepatic bile salt clearance was only marginally abnormal in one subject. The presence of hyperlipidaemia does not invalidate the use of serum conjugated bile salt analysis for the detection of liver diseases.

The synthesis and characterisation of a novel iodinated bile salt derivative, 125I-labelled 3 beta-azidocholylhistamine, is described. The derivative is handled by rat liver in a similar manner to taurocholate and binding to bovine serum albumin, a well-characterised bile acid-binding protein, is demonstrated. The suitability of the derivative for photoaffinity labelling is assessed.

The concentrations of conjugated chenodeoxycholate and cholate in serum have been measured in 26 controls and 19 patients with liver disease before and after taking a fat-containing meal. The times at which maximum bile salt concentrations occurred varied considerably between individuals. In the majority of cases maximal concentrations of the two bile salts occurred simultaneously but in 3 subjects maximal concentrations of chenodeoxycholate were found 15-60 min prior to maximal cholate concentrations. The measurement of bile salt concentration in a single sample taken 2 h after the fatty test meal would have produced two false-negative results in the patients with liver disease. It is concluded that the jejunal absorption of dihydroxylated bile salts may not be significant in the majority of individuals and that the protocol for assessment of liver function by postprandial bile salt analysis should include at least two blood samples collected at 1.5 and 2 h after ingestion of the test meal.

1. A compartmental model has been used to derive the in vivo subcellular distribution of lithocholic acid in rat liver. The model is based on the values of the partition coefficients for the distribution of lithocholic acid between subcellular fractions and buffer. It also permits calculation of the amount of lithocholic acid which is in free solution in cytosol. 2. The hypothesis that the rate of biliary excretion of a bile acid depends on the proportion in free solution was investigated by comparing the rates of biliary excretion of lithocholic acid and glycocholic acid. The rate for lithocholic acid was substantially less than for glycocholic acid while the percentages of each bile acid in free solution were 0.8% and 10%, respectively. 3. The validity of the model was supported by the observation that the amounts of lithocholic acid predicted to be present in the nuclear and cytosolic fractions were similar to the amounts found after intravenous injection of the bile acid.

The clearance of an injected dose of sodium glycocholate has been studied in a group of 23 control subjects who had no evidence of liver disease. A significant correlation was found between bile salt clearance and age. No significant difference was found between the bile salt clearance of male and female subjects. Data derived from a simple compartmental model indicated that decreased clearance of bile salt with age was not specifically related to degenerative changes in hepatocellular integrity, hepatic uptake of bile salts or bile flow.

1. Two lithocholic acid-binding proteins in rat liver cytosol, previously shown to have glutathione S-transferase activity, were resolved by CM-Sephadex chromatography. 2. Phenobarbitone administration resulted in induction of both binding proteins. 3. The two proteins had distinct subunit compositions indicating that they are dimers with mol.wts. 44 000 and 47 000. 4. The two lithocholic acid-binding proteins were identified by comparing their elution volumes from CM-Sephadex with those of purified ligandin and glutathione S-transferase B prepared by published procedures. Ligandin and glutathione S-transferase B were eluted separately, as single peaks of enzyme activity, at volumes equivalent to the two lithocholic acid-binding proteins. 5. Peptide 'mapping' revealed structural differences between the two proteins.

A scheme for kit evaluation in the individual clinical chemistry laboratory is presented. Recommendations for assessing accuracy, precision, and stability are included along with a scheme for preparing for the evaluation and writing the final report.

Investigations into the choice of immunogen, ligand, antiserum and assay conditions for the radioimmunoassay of conjugated cholic acid have been performed with a view to producing optimal assay conditions. Cholic acid-BSA was found to be the best immunogen to produce antibodies to conjugated cholic acid and the response was of an IgG type. Incorporating a spacer (hexanoic acid) between hapten and carrier protein resulted in a decrease in antiserum titre. Optimal conditions for the assay were found using [125I]histamine-glycocholic acid as ligand with a dilution of antiserum to produce 60% binding of ligand and a pH of 7.4. Using these assay conditions no serum effects were found; extraction of serum prior to assay was therefore unnecessary. The assay was sensitive enough to detect post-prandial increased in serum bile acid concentrations following a liquid test meal; no increase was observed throughout the same time period in a fasting control.

The peak-monitor software of the Technicon SMAC system has been investigated by experiments designed to test its ability to detect abnormally shaped peaks, deliberately produced by presenting serum samples of inadequate volume for analysis. The software provided by the manufacturer is shown to detect faulty peaks inefficiently, and a set of modified parameters has been identified which greatly improves the performance of the peak-monitor software.

1. The partial purification of two lithocholic acid-binding proteins from liver 100 000g supernatants is described. 2. Gel-filtration, (NH4)2SO4 fractionation, Ca3(PO4)2 fractionation and ion-exchange chromatography were used. 3. Both proteins exhibited glutathione S-transferase activity; one may be the non-specific anion-binding protein ligandin. 4. Glutathione S-transferase activity of one of the binding proteins was inhibited by lithocholic acid.