Administration Of BM Research Articles

Discrimination of Vitis vinifera varieties using DNA fingerprinting and NMR coupled with chemometrics and their impact on the efficacy of fluoxetine and indomethacin in vivo

The grapes, Vitis vinifera, belong to family Vitaceae. Some fruit juices substantially decrease the oral absorption of certain drugs undergoing intestinal uptake transport. Green seedless “Thompson” type (GT) and reddish black seeded “Magic black” type (BM) of V. vinifera L. were discriminated using deoxyribonucleic acid (DNA) fingerprinting and nuclear magnetic resonance (NMR) coupled with chemometrics. Their effects on the bioavailability of fluoxetine and indomethacin were assessed in vivo. Practical applications Random amplification of polymorphic DNA (RAPD) analysis of GT and BM revealed that primers OPA-02 and OPB-10 indicate a taxonomical relationship between them as they produced high similarity coefficients estimated by 80 and 66.67%, respectively. OPC-03, OPB-13, and OPC-07 primers help in their discrimination based on their minimal levels of similarity coefficients estimated by zero percentages accompanied by the high levels of polymorphism. Principal component analysis score plot obtained from processing of proton nuclear magnetic resonance (1H NMR) showed very good discrimination between GT and BM in PC1 that accounts for most of the variance. Oral administration of 100 mg/kg of GT and BM separately produce no significant change in immobility period of rats dropped individually into a glass cylinder. Concomitant administration of either 100 mg/kg of GT or BM with 20 mg/kg of fluoxetine significantly decreased the efficacy of antidepressant reference drug (fluoxetine) by decreasing time before immobility showing a percentage of depression inhibition estimated by 17.2 and 29.1%, respectively, that obviously decreased when compared to fluoxetine alone. Regarding anti-inflammatory activity, both GT and BM exhibited a notable activity by inhibiting the increase in paw edema and inflammation induced by carrageen in rats where BM is more potent approaching indomethacin. Administration of GT or BM with indomethacin reduces its efficacy revealing 27.5 and 26.1% in edema percentage after 1 hr, respectively, that decreased minimally to be 19.1% and 15.6% after 4 hr, respectively, instead of 7% in case of indomethacin alone. Thus, it can be concluded that DNA and NMR fingerprinting could successfully differentiate between Vitis varieties. Grape juice decreased the efficacy of fluoxetine and indomethacin. Further studies should be conducted to declare the effect of Vitis varieties on the bioavailability of additional drugs that treat chronic diseases as diabetes and hypertension.

MSCs ameliorates DPN induced cellular pathology via [Ca2+ ]i homeostasis and scavenging the pro-inflammatory cytokines.

The MSCs of various origins are known to ameliorate or modulate cell survival strategies. We investigated, whether UCB MSCs could improve the survival of the human neuronal cells and/or fibroblast assaulted with DPN sera. The results showed, the co-culture of UCB MSCs with human neuronal cells and/or fibroblasts could effectively scavenge the pro-inflammatory cytokines TNF-α, IL-1β, IFN-ɤ and IL - 12 and control the pro-apoptotic expression of p53/Bax. Further co-culture of UCB MSCs have shown to induce anti-inflammatory cytokines like IL-4, IL-10 and TGF-β and anti-apoptotic Bclxl/Bcl2 expression in the DPN sera stressed cells. Amelioration of elevated [Ca2+ ]i and cROS, the portent behind the NFκB/Caspase-3 mediated inflammation in DPN rescued the cells from apoptosis. The results of systemic administration of BM MSCs improved DPN pathology in rat as extrapolated from human cell model. The BM MSCs ameliorated prolonged distal motor latency (control: 0.70 ± 0.06, DPN: 1.29 ± 0.13 m/s DPN + BM MSCs: 0.89 ± 0.02 m/s, p < 0.05) and lowered high amplitude of compound muscle action potentials (CMAPs) (control: 12.36 ± 0.41, DPN: 7.52 ± 0.61 mV, DPN + MSCs: 8.79 ± 0.53 mV, p < 0.05), while slowly restoring the plasma glucose levels. Together, all these results showed that administration of BM or UCB MSCs improved the DPN via ameliorating pro-inflammatory cytokine signaling and [Ca2+ ]i homeostasis.

Antenatal glucocorticoids reduce growth in appropriately grown and growth-restricted ovine fetuses in a sex-specific manner

Antenatal glucocorticoids are administered to mature the fetal lungs before preterm birth. Glucocorticoids also have non-pulmonary effects, including reducing fetal body and brain growth. The present study examined whether glucocorticoid administration has a sex-specific effect on growth in appropriately grown (control) and intrauterine growth-restricted (IUGR) fetal sheep. IUGR was induced at 0.7 gestation in fetal sheep by single umbilical artery ligation. On Days 5 and 6 after surgery, IUGR or control fetuses were exposed to the synthetic glucocorticoid betamethasone (BM; 11.4mg) or saline via intramuscular maternal administration. On Day 7, a postmortem was conducted to determine fetal sex and weight. Compared with control fetuses, the birthweight of male and female IUGR fetuses was significantly reduced (by 18.5±4.4% (P=0.002) and 21.7±6.0% (P=0.001), respectively). Maternal administration of BM significantly reduced bodyweight in both control and IUGR fetuses (by 11.3±2.8% and 20.5±3.6% in control male and female fetuses, respectively; and by 22.9±3.1% and 38.3±3.4% in IUGR male and female fetuses, respectively; P<0.001 for all, versus control+saline) fetuses. In control and IUGR animals the degree of growth restriction was greater in females than males (P<0.05) following administration of BM. These data suggest that antenatal glucocorticoids reduce fetal growth in a sex-specific manner, with females more growth restricted than males.

Strategies to enhance T cell reconstitution following severe immunodepletion

Allogeneic hematopoietic stem cell transplant (AHSCT) is one of the only curative therapies for leukemia patients and others with hematological malignancies. Its use is restricted by major complications including post‐transplant immune deficiency which is devastating to the T cell compartment. Promotion of rapid and effective reconstitution of T cell immunity reduces incidence of transplant‐related complications and significantly improves the survival of HSCT recipients. Our previous studies show that precursor T cells (preT) generated ex vivo from stem cells on the OP9‐DL1 culture system can significantly enhance T cell reconstitution in AHSCT recipients. However, despite their regenerative benefits, the homing and seeding of preT to the thymus is relatively inefficient (only 1 out of every 10,000 iv injected preT engraft in the thymus). In the present study we examined strategies to improve the effectiveness of preT treatment. We found that direct administration of lineage‐depleted BM into the thymus can have significant benefit for T cell reconstitution. We also found that small changes in the purification strategy for seeding OP9‐DL1 cultures (based on differential CD150 expression) can have significant impacts on the generation and output of preT. These findings lay the foundation for improving T cell rejuvenation by enhancing the utility of preT as a potential clinical therapy.

Effects of GABA receptor blockade on the ventilatory response to hypoxia in hypothermic newborn piglets.

Hypothermic newborn piglets have a depressed ventilatory response to hypoxia, and this may be due to an increase in CNS gamma-aminobutyric acid (GABA) levels. To evaluate the effects of GABA(A) receptor blockade on the ventilatory response to hypoxia in hypothermic piglets, 31 anesthetized paralyzed mechanically ventilated newborn piglets (2-7 d) were studied at a brain temperature of 38.5 +/- 0.5 degrees C [normothermia (NT), n = 15] or 34 +/- 0.5 degrees C [hypothermia (HT), n = 16]. The central respiratory output was evaluated by measuring burst frequency and moving time average area of phrenic nerve activity. Measurements of minute phrenic output (MPO), arterial blood pressure, heart rate, oxygen consumption, and arterial blood gases were obtained at room air and during 20 min of isocapnic hypoxia [fraction of expired oxygen (FiO2) = 0.10]. After 10 min of hypoxia, a bolus injection of 20 microL of bicuculline methiodide (BM; 10 microg) or Ringer's solution was administered into the cisterna magna over a 1-min period, and the piglets remained in hypoxia for an additional 10 min. There was an initial increase of 50 +/- 6% in MPO during the first minute of hypoxia followed by a decrease to values 24 +/- 8% above baseline at 10 min in the NT group. In contrast, in the HT group, the initial increase in MPO with hypoxia was eliminated, and, at 10 min, there was a decrease to a mean value 35 +/- 4% below baseline level (NT versus HT, p < 0.03). After administration of BM, a significant increase in MPO with hypoxia was observed in both groups compared with their placebo groups (p < 0.002 in NT-BM group, p < 0.0001 in HT-BM group). However, the magnitude of the increase in MPO during hypoxia was significantly greater in the HT group after administration of BM (NT versus HT, p < 0.0001). Changes in oxygen consumption, arterial blood pressure, heart rate, pH, partial pressure of oxygen (PaO2), and base excess with hypoxia were not different between NT and HT groups before and after the administration of BM. The cardiorespiratory response to hypoxia was not modified after administration of Ringer's solution to NT and HT placebo groups. These data suggest that the depression in hypoxic ventilatory response produced by HT is in part modulated by an increased CNS GABA concentration.

Plasma copper and iron changes in sheep after left lung inhalation injury: effect of the thromboxane antagonist BM 13.177 (Solutroban).

A significant decline in plasma concentrations of copper and iron were observed in sheep exposed to preferential smoke inhalation of the left lung. The decline was evident 30 minutes after smoke inhalation, and the levels of both trace metals persisted at quite low levels for up to the 18-hour time interval after injury. From that time a gradual recover for copper but not for iron levels was observed so that by 24 hours the levels of copper were in the same range of those at baseline. Copper and iron levels showed an inverse correlation to airway peak and plateau pressures and left lung vascular resistance index and a direct correlation to left lung blood flow. Administration of BM 13.177 (Solutroban), a thromboxane antagonist, before exposure to smoke inhalation protected the sheep from the decline of copper and iron levels in plasma. In these animals airway peak and plateau pressure, left lung vascular resistance, and blood flow were also unmodified. Lipid peroxidation of the lung tissue by oxygen free radicals were lower than in those animals that did not receive BM 13.177. There was likewise a tendency of a decreased wet-to-dry weight ratios in the animals treated with BM 13.177. BM 13.177 treatment in an inhalation injury model might partly protect lung damage and parallels unchanged plasma copper and iron levels. The plasma copper and iron may therefore be an indicator of acute lung damage.

Endothelin-1-induced myocardial ischaemia and oedema in the rat: involvement of the ETA receptor, platelet-activating factor and thromboxane A2.

1. The objectives of the present experiments were to assess the role of ETA receptors in mediating endothelin-1 (ET-1)-induced myocardial ischaemia and oedema and to study the involvement of platelet-activating factor (PAF) and thromboxane A2 (TxA2) in these actions of ET-1 in rats. 2. Intravenous bolus injection of ET-1 (0.1-2 nmol kg-1) into anaesthetized rats induced ST segment elevation of the electrocardiogram in a dose-dependent manner without causing arrhythmias. ST segment elevation developed within 20-90 s and persisted for at least 10-20 min following administration of ET-1. 3. Pretreatment of the animals with the selective endothelin ETA receptor antagonist, FR 139317 (2.5 mg kg-1, i.v.) inhibited by 86% the ST segment elevation elicited by ET-1 (1 nmol kg-1). Pretreatment with intravenous administration of BM 13505 (1 mg kg-1), a TxA2 receptor antagonist, OKY-046 (10 mg kg-1), a thromboxane synthase inhibitor or the specific PAF receptor antagonist, WEB 2086 (1 mg kg-1) or BN 52021 (10 mg kg-1) markedly suppressed ST segment elevation in response to ET-1. Infusion of indomethacin (3 mg kg-1 bolus plus 2 mg kg-1 h-1) did not significantly affect ET-1-induced ST segment elevation. 4. Bolus injection of ET-1 (1 nmol kg-1, i.v.) to conscious rats resulted in a prolonged pressor effect preceded by a transient depressor response. Corresponding to changes in blood pressure, a small transient tachycardia was followed by a sustained bradycardia. ET-l enhanced albumin leakage by 87 and 120% in the left ventricle and right atrium, respectively, as measured by the extravasation of Evans blue dye.5. The selective ETA receptor antagonist, FR 139317 (2.5 mg kg-1) significantly blunted the pressor action of ET-1 and the accompanying bradycardia without affecting the depressor response. Furthermore,FR 139317 almost completely abolished the permeability effect of ET-l in both vascular beds studied.6. Pretreatment of the animals with BM 13505 (1 mg kg-1), OKY-046 (10mg kg-1), WEB 2086(1 mg kg-1) or BN 52021 (10mg kg-1) significantly reduced ET-1 (1 nmol kg-1)-induced albumin extravasation both in the left ventricle and right atrium. The PAF receptor antagonists, WEB 2086 and BN 52021 were equally potent inhibitors in the left ventricle, whereas BN 52021 appeared to be a more potent inhibitor than WEB 2086 in the right atrium. Pretreatment with indomethacin (3 mg kg-1 plus 2 mg kg-1 h-1) did not modify the permeability response to ET-1. None of these compounds affected significantly ET-l-induced changes in mean arterial blood pressure and heart rate.7. These results indicate that intravenous administration of ET-1 provokes ST segment elevation and myocardial oedema and suggest that these events are mediated, in part, through release of secondary mediators, such as PAF and TxA2 via the activation of ETA receptors.

Induction of the major integral membrane protein of mouse liver peroxisomes by peroxisome proliferators.

Peroxisome proliferators are known to increase the volume of the peroxisomal compartment in rodent liver. We have examined the induction of the major integral membrane protein of mouse liver peroxisomes (PMP68) by a number of these agents, and compared this with their effect on the peroxisomal bifunctional protein (PBP), an enzyme of the beta-oxidation pathway which is located in the peroxisome matrix. Dietary clofibrate, di-2-(ethylhexyl)phthalate and Wy-14,643, three structurally unrelated proliferators, all increased the mRNA and protein content of PMP68 approx. 2-fold, whereas PBP was induced 8-13-fold. The kinetics and sequence of induction of PMP68 and PBP following a single dose of Wy-14,643 were compared and shown to be similar, and the effects were reversible. Another proliferator, BM 15766, caused maximal induction of PMP68 but only a low induction of PBP; further PBP induction was achieved by the administration of BM 15766 in combination with Wy-14,643. Similarly, BM 15766 and Wy-14,643 increased transcription of the PMP68 gene in vitro, whereas PBP gene transcription was increased by Wy-14,643 but not by BM 15766. Thus peroxisome proliferators enhance the expression of the genes for both the membrane protein PMP68 and the matrix protein PBP, but the regulation of this expression appears to be mediated by different mechanisms.

Diets Enriched With N‐3 Fatty Acids Ameliorate Lactic Acidosis by Improving Endotoxin‐Induced Tissue Hypoperfusion in Guinea Pigs J.J. POMPOSELLI, E.A. FLORES, G.L. BLACKBURN, S.H. ZEISEL, B.R. BISTRIAN Ann Surg 213:166–176, 1991

The authors compared the effects of 6 weeks of dietary supplementation with either N‐3 fatty acids (fish oil) or N‐6 fatty acids (safflower oil) on the acute physiologic response to continuous endotoxin infusion in guinea pigs. Substantial evidence has accumulated to suggest that arachidonic acid metabolites may act as a common efferent signal in the pathogenesis of septic shock. Studies attempting to elucidate the physiologic effects of diets rich in N‐3 fatty acids have centered on the altered formation and metabolism of prostanoids. Eicosapentaenoic acid (EPA) has been shown to be the metabolically active fatty acid in fish oil because of its structural similarity to the prostaglandin precursor, arachidonic acid (AA). Controlled dietary intake of selected fish oils rich in EPA and N‐3 fatty acids have been shown to down‐regulate eicosanoid metabolism leading to a more vasodilatory platelet and neutrophil antiaggregatory state compared with diets enriched with N‐6 fatty acids. In previous studies, the authors have shown that dietary pretreatment with N‐3 fatty acids attenuated the febrile response, reduced thromboxane formation to exogenous IL‐1 administration, and improved survival after endotoxin challenge in guinea pigs. In the present study, it is hypothesized that diets enriched with N‐3 fatty acids may alter the physiologic response to endotoxin, possibly by altering membrane phospholipids.Three different experimental protocols were carried out after a 6‐week feeding period in which animals received a standard diet containing either menhaden fish oil (N‐3 fatty acids) or safflower oil (N‐6 fatty acids). In the first experiment, animals were infused with endotoxin for 7 hours and blood pH, paCO2, bicarbonate, blood pressure, plasma lactate, serum phospholipid fatty acid profile, and histology (lung, liver, and kidney) were examined. In the second experiment, cardiac output and blood flow distribution during endotoxemia were determined. In the third experiment, the effect of either indomethacin or a specific thromboxane A2 receptor blocker (BM 13505) on the response to endotoxemia was examined. An additional subset of animals were infused with dichloroacetic acid (DCA), which prevents inhibition of the pyruvate dehydrogenase enzyme complex by endotoxin.The mortality rate of animals infused with endotoxin alone was 20% in the safflower oil‐fed group and 10% in the menhaden oil‐fed group (not a statistically significant difference). No deaths were observed in animals pretreated with either indomethacin or BM 13505 regardless of diet. Safflower oil‐fed animals developed an initial respiratory alkalosis followed by metabolic acidosis in response to endotoxin infusion, whereas fish‐oil fed animals developed a mild respiratory alkalosis and no metabolic acidosis. Pre‐treatment with indomethacin or BM 13505 had similar effects to N‐3 fatty acid feeding in ameliorating the development of metabolic acidosis in safflower oil‐fed animals, suggesting that prostaglandins and especially thromboxane, may be responsible for the development of some of the sequelae of endotoxemia. Safflower oil‐fed animals had a sevenfold increase in lactate production compared with controls and a threefold increase compared with fish oil‐fed animals. Infusion of indomethacin, significantly reduced the amount of lactate formed in safflower oil‐fed animals given endotoxin, but had no additive effect in fish oil‐fed animals. Administration of BM 13505 was only partially effective in reducing lactate levels, which remained significantly above control levels. The infusion of DCA failed to lower blood lactate levels in both groups given endotoxin. Safflower oil‐fed animals developed a diffuse interstitial pneumonia with increased neutrophils and intravascular congestion. Fish oil‐fed animals infused with endotoxin demonstrated intravascular congestion in their lungs without infiltrates. Examination of other tissues did not demonstrate any abnormalities in all groups, with the exception of the liver, which showed mild fatty infiltration in both dietary groups. Lung function was not severely compromised during the initial period of endotoxemia in either group. Endotoxin infusion caused a significant decrease in percent perfusion of both skin and muscle, regardless of dietary intake. However, in muscle, decreased perfusion was partly prevented by pre‐treatment with N‐3 fatty acids.

Reduction of Myocardial Damage and Polymorphonuclear Leukocyte Accumulation Following Coronary Artery Occlusion and Reperfusion by the Thromboxane Receptor Antagonist BM 13.505

This study was designed to assess the effect of the thromboxane receptor antagonist, BM 13.505, in limiting myocardial damage and polymorphonuclear leukocyte accumulation in rats subjected to coronary artery occlusion for 30 min with reperfusion for 24 h (MI/R). Myocardial injury and polymorphonuclear leukocyte infiltration were determined by measuring creatine phosphokinase (CPK) specific activity and myeloperoxidase (MPO) activity, respectively, in the left ventricular free wall (LVFW). Myocardial CPK levels were 8.24 +/- 0.33 U/mg protein in sham MI/R-vehicle-treated animals (n = 18), and were significantly decreased to 6.51 +/- 0.44 U/mg protein in MI/R-vehicle animals (n = 22). Myocardial MPO values were 2.4 +/- 0.5 U/g LVFW in sham MI/R animals, and significantly increased to 10.9 +/- 1.3 U/g LVFW in MI/R-vehicle animals. Administration of BM 13.505 (30 mg/kg, i.p.) 1 min prior to coronary occlusion resulted in CPK values of 7.83 +/- 0.45 U/mg protein and MPO levels of 6.1 +/- 0.9 U/g LVFW (p less than 0.05, compared to the MI/R-vehicle group). The survival rate in the MI/R-BM 13.505 group was 74 and 65% at 2 and 24 h, respectively, and was not different from the MI/R-vehicle group. There were no significant differences in mean arterial blood pressure or heart rate between the MI/R-vehicle and MI/R-BM 13.505 groups, indicating that changes in myocardial oxygen demand do not explain the protective effects. A lower dose did not reduce myocardial injury, indicating that the effects of BM 13.505 were dose dependent.(ABSTRACT TRUNCATED AT 250 WORDS)

Protective effects of the thromboxane receptor antagonists BM 13.177 and BM 13.505 against U 46619-induced sudden death in rats.

The purpose of this study was to evaluate the pharmacology and pharmacodynamics of the thromboxane receptor antagonists, BM 13.177 and BM 13.505, for prevention of U 46619-induced sudden death in anesthetized male Sprague-Dawley rats. The injection of U 46619 (100 micrograms/kg i.v.) produced sudden death typically between 5 and 15 min. Administration of 0.01 mg/kg BM 13.505 (i.v.) 0.1 h prior to the U 46619 challenge did not protect against sudden death, while doses of 0.03 mg/kg or greater protected completely (100% survival). A dose of 1 mg/kg BM 13.505 afforded protection to 2 h but not to 24 h, while a single dose of 30 mg/kg administered 24 h prior to the U 46619 challenge provided complete protection against the lethal event. Administration of BM 13.177 (30 mg/kg, i.v. or i.p.) blocked the effects of U 46619 when administered 0.1 h before the challenge, but not when given 2 or 3 h prior to the challenge with U 46619. Pretreatment with indomethacin did not block the effects of U 46619, indicating that formation of endogenous thromboxane does not play a major role in the lethal effects of U 46619, and that blockade of the lethal effects of U 46619 was specific for thromboxane receptor antagonists. These data demonstrate that BM 13.177 and BM 13.505 prevented sudden death produced by the injection of U 46619. At comparable doses of 30 mg/kg, the duration of action for BM 13.505 was was significantly greater than for BM 13.177. These data suggest that BM 13.177 and BM 13.505 may be useful for the investigation of diseases where thromboxane is involved.

Stimulation by low phosphorus and low calcium diets of duodenal absorption of phosphate in betamethasone-treated chicks.

Although the inhibitory effect of glucocorticoids on the intestinal absorption of calcium is well recognized, their effect on the absorption of phosphate is less well documented. We studied the effect of the oral administration of betamethasone (BM; 25 micrograms/kg per day) on the duodenal absorption of phosphate in chicks fed normal calcium, normal phosphorus (NCaNP), normal calcium, low phosphorus (NCaLP) or low calcium, normal phosphorus (LCaNP) diets using the ligated loop technique in vivo. The daily oral administration of BM for 8 days significantly reduced the absorption of phosphate in chicks fed the NCaNP diet (21% decrease) but had less effect in chicks fed the NCaLP (14% decrease) or LCaNP (9% decrease) diets in which birds the absorption of phosphate was significantly raised (49 and 87% respectively). In one group of chicks, BM was administered for 9 days before the birds were transferred to the NCaLP or LCaNP diets. Adaptation was again unaffected by the treatment. Thirty-four per cent of the absorbed phosphate was retained in the duodenal tissue. Treatment with BM reduced the amount retained but this may have been caused by the lower weight of the duodenal segment in these chicks as BM administration markedly reduced growth rate. We have concluded that the duodenal absorption of phosphate in the chick can be inhibited by treatment with BM, although this may be secondary to the reduced rate of growth, but the increase in the absorption of phosphate caused by feeding NCaLP or LCaNP diets was unaffected by the steroid.

Therapeutic effects of BM 12,531 (Prop. INN azimexon).

The antitumor effect of BM 12,531, 2-[2-cyanaziridinyl-(1)]-2[carbamoylaziridinyl-(1)]-propane (Prop. INN azimexon), on L1210 leukemia in mice is described. The immunostimulating compound BM 12,531 can also reduce the acute toxicity of X-irradiation in mice. The most effective doses in these experiments are 1 or 5 mg/kg twice a week, whereas 50 and 500 mg/kg twice a week decrease the therapeutic effect. The mortality of Candida-infected and irradiated mice is also reduced by oral administration of BM 12,531. BM 12,531 has synergistic effects with subtherapeutic doses of chloramphenicol in an experimental E. coli infection in mice.

Radioimmunoassay for plasma betamethasone and its application for clinical study (author's transl)

A sensitive and specific radioimmunoassay for plasma betamethasone has been developed. The antiserum used was obtained by immunizing rabbits with betamethasone-3-oxime-human serum albumin (BM-3-oxime-HSA) conjugate. The sensitivity was 20 pg and a standard curve was established with a useful range from 20 pg to 4 ng. The cross reactivity of all tested endogenous steroids was less than 0.2%. Cortisol with a cross reactivity of 0.14% caused slight interference at very high concentrations, but this factor is negligible when plasma cortisol level is less than 30 microgram/dl. The calculated interference by cortisol was less than 10%, and plasma not receiving BM consistently gave blanks which were less than 20 pg/tube. Reliability criteria were satisfactory. By this method plasma BM could be measured directly in dichloromethane extract of plasma. The plasma concentrations of BM were measured in normal subjects and patients with liver diseases following oral administration of BM. The peaks of the plasma concentrations for 1.0 mg and 1.5 mg of BM were 345 +/- 40 (n = 3) and 710 +/- 200 ng/dl (n = 5) respectively within 2 hours after administration in normal subjects. After the peak level, plasma BM rapidly fell and disappeared 24 hours after administration in all examined normal subjects. In patients with chronic active hepatitis, the peak levels for 1.0 mg and 1.5 mg of BM were 428 +/- 48 (n = 4) and 837 +/- 83 ng/dl (n = 7) respectively within 2 hours after administration. However, the peak levels of plasma BM were higher than those of normal subjects, and the disappearance of BM from the blood was markedly delayed, reaching a level of 318 +/- 88 nad 622 +/- 148 ng/dl respectively for 1.0 mg and 1.5 mg of BM at 5 hours. The relatively high plasma concentration of BM, ranging from 135 to 170 ng/dl was maintained even 24 hours after administration in all patients with chronic active hepatitis. The disappearance of cortisol from the blood also rapidly fell in normal subjects, but was markedly delayed in patients with chronic active hepatitis. There was good correlation between the severity of the liver disease as measured by the ICG retention at fifteen minutes and removals of BM and cortisol from the blood.