Enzyme activities in tissues and elimination half-lives of homologous muscle and liver enzymes in the racing pigeon (Columba Livia domestica)

Tissue enzyme profiles of heart, liver, pectoral muscle, quadriceps muscle, duodenum, kidney and brain from racing pigeons were established. The enzymes were alanine aminotransferase (ALAT), aspartate aminotransferase (ASAT), glutamate dehydrogenase (GLDH), lactate dehydrogenase (LDH), gamma glutamyltransferase (y-GT), alkaline phosphatase (AP), and creatine kinase (CK). Elimination half-lives (tyss) of certain enzymes were also determined. The mean values (+/- SD) were: ASAT, 7.66 +/- 1.55 (liver) and 6.51 +/- 0.83 (muscle); ALAT, 15.69 + 1.70 (liver) and 11.99 +/- 1.32 (muscle); LDH, 0.71 + 0.10 (liver) and 0.48 +/- 0.07 (muscle); GLDH, 0.68 +/- 0.17 (liver), CK 3.07 +/- 0.59 hours (muscle). GLDH is the most liver-specific enzyme in the pigeon, but increased activities in the plasma are likely only in the acute stage of severe liver cell damage, since this enzyme is localised within the mitochondria and has a short half life. LDH and ASAT seem to be the most sensitive indicators of liver cell damage, though contributions come from muscle damage. Muscle cell damage can be differentiated from liver cell damage by measuring plasma CK activity, since CK is both a specific and a sensitive indicator of muscle cell damage. In a clinical setting the combined use of LDH, ASAT and CK permits differentiation between liver and muscle cell damage in racing pigeons.     

Effects of the prostaglandin E2 analogue enprostil on the carbon tetrachloride-induced necrosis of liver cells in mice.

Female mice, eight weeks old, were injected with carbon tetrachloride (CCl4) (10 mg subcutaneously). Groups of mice (n = 10-30) were then injected with enprostil (E) 2, 20 or 50 micrograms/kg body weight (bw) intraperitoneally 15 min and two h after, or E 100 micrograms/kg bw two h after the CCl4 injection. The mice were killed after 24, 48 or 72 h. Plasma activity concentrations of alanine aminotransferase (ALAT) were determined in blood specimens from the iliac veins. The extent of liver cell necrosis in histological sections was recorded on a 100 mm Visual Analogue Scale (VAS) and measured using the electronic Mini Mop method. In the group given the highest single dose of E (100 micrograms/kg) a significant lowering of the CCl4-induced liver cell necrosis was found after 24 h. No significant differences were found after 48 and 72 h. In the other groups injected with lower doses of E after CCl4, no significant differences were found compared to groups injected with CCl4 alone.     

Changes in plasma chemistry after drug-induced liver disease or muscle necrosis in racing pigeons (Columba livia domestica)

Changes in plasma variables as a result of liver damage induced by ethylene glycol (group A) or D-galactosamine (group B) and of muscle damage induced by doxycycline were compared. Plasma bile acid concentration was both a specific and a sensitive indicator of liver disease. Another specific, but less sensitive indicator of liver disease was 7-GT. Plasma AS AT activity was the most sensitive indicator of disease of the liver, but was not specific, since increased ASAT activities were also seen during muscle disease. ALAT activity was slightly more sensitive to liver damage than 7-GT, but was also not specific, being increased also after muscle damage. Plasma GLDH activity was increased only as a result of extensive liver necrosis. AP activity was of no value for detecting liver disease in the pigeon. CK activity was specific for muscle injury, though the activities of ALAT, ASAT and LD were also increased. Because of its long elimination half-life, increased ALAT activity persisted for 9 days after muscle damage, whereas CK activity returned to reference values within 3 days. LDH was a poor indicator of damage to liver and muscle, despite its relatively high tissue concentrations in both tissues. The rapid disappearance rate of LDH from plasma probably explains this observation.     

Cellular Cl content and concentration of amphibian skeletal and heart muscle.

Toads (Bufo marinus) and frogs (Rana pipiens pipiens) were given intraperitoneal injections of Na36Cl and Na235SO4. After in vivo equilibration for 20--180 min, the animals were pithed, and their ventricular and semitendinosus muscles were excised. Measurements of total Cl (by titrimetry) and 36Cl (by radioassay) showed that specific radioactivities of plasma and mus les approached equality within 1 h after injection for toad skeletal and heart muscle and frog ventricles, indicating complete exchange of cellular Cl with 36Cl. From the simultaneously measured muscle water contents and 35SO4 spaces, intracellular Cl concentrations in vivo (in mumol/g cell water) for semitendinosus and ventricular muscles were calculated to be, respectively, 1.4 +/- 0.3 and 2.3 +/- 0.8 for Bufo and 1.7 +/- 0.7 and 4.8 +/- 2.4 for Rana. In view of these low values, active cellular Cl accumulation seems improbable, but cannot be rigorously excluded without simultaneous membrane potential and intracellular ion activity measurements. A high concentration of Cl in the sarcoplasmic reticulum of skeletal muscle is also inconsistent with these measurements.     

Plasma levels of somatostatin following a test meal in dogs. Initial atropine resistant vagally mediated decrease of somatostatin levels and increase of gastrin and insulin levels

Plasma levels of somatostatin like immunoreactivity (SLI), below referred to as somatostatin levels, were measured in peripheral plasma of conscious dogs. Basal somatostatin levels averaged 49 +/- 10 pM. Somatostatin as well as gastrin and insulin plasma levels were measured before and after feeding with and without prior atropinization. During the first 10 min after feeding somatostatin levels fell from 49 +/- 10 to 23 +/- 9 pM, whereas gastrin and insulin levels rose from 9 +/- 2 and 140 +/- 14 pM to 48 +/- 11 and 370 +/- 91 pM respectively. Atropine 0.01 or 0.1 mg/kg did not inhibit these responses. After the initial decrease, somatostatin level rose again and peaked at around 60 min after feeding (110 +/- 24 pM). This secondary rise was completely abolished by atropine in both doses tried. Gastrin and insulin levels remained elevated throughout the experiments with and without atropine. It is suggested that gastrin release and HCl secretion are inhibited by a tonic outflow of gastric somatostatin during basal conditions. The process of feeding induces an atropine resistant, vagally mediated decrease in somatostatin release from the gastrointestinal tract and this decreased output of somatostatin facilitates initiation of meal-related endocrine and exocrine gastric secretions.     

The effect of carbon tetrachloride on the copper-laden rat liver

Copper is believed to be hepatotoxic in Indian Childhood Cirrhosis and Wilson's disease. However, copper-loading causes only minimal hepatic damage in animal models. The hypothesis was therefore proposed that a second hepatic insult may precipitate or perpetuate liver injury in a copper-laden liver. In non-copper-dosed rats CCl4 (10 mmol/kg, i.p.) produced elevated serum AST (809 +/- 298 IU/l, normal 20 +/- 5) and ALT (295 +/- 157 IU/l, normal 6 +/- 1) and extensive liver cell necrosis, portal tract inflammation, fat deposition, and perilobular hepatocyte ballooning. In rats whose liver copper was elevated from 75 +/- 13 to 461 +/- 13 micrograms/g by oral copper supplementation, CCl4 produced much smaller increases in AST (492 +/- 80 IU/l) and ALT (172 +/- 57 IU/l) and mild focal liver cell necrosis. Fat deposition and perilobular vacuolation were not reduced. Prior copper-loading of rats unequivocally protected against the CCl4-induced liver injury. Triglyceride accumulation, however, was apparently unaffected. The possible interactions of copper with prostaglandin-mediated inflammation and with free-radical-induced liver damage are discussed.     

The effect of carbon tetrachloride on the copper-laden rat liver.

Copper is believed to be hepatotoxic in Indian Childhood Cirrhosis and Wilson''s disease. However, copper-loading causes only minimal hepatic damage in animal models. The hypothesis was therefore proposed that a second hepatic insult may precipitate or perpetuate liver injury in a copper-laden liver. In non-copper-dosed rats CCl4 (10 mmol/kg, i.p.) produced elevated serum AST (809 +/- 298 IU/l, normal 20 +/- 5) and ALT (295 +/- 157 IU/l, normal 6 +/- 1) and extensive liver cell necrosis, portal tract inflammation, fat deposition, and perilobular hepatocyte ballooning. In rats whose liver copper was elevated from 75 +/- 13 to 461 +/- 13 micrograms/g by oral copper supplementation, CCl4 produced much smaller increases in AST (492 +/- 80 IU/l) and ALT (172 +/- 57 IU/l) and mild focal liver cell necrosis. Fat deposition and perilobular vacuolation were not reduced. Prior copper-loading of rats unequivocally protected against the CCl4-induced liver injury. Triglyceride accumulation, however, was apparently unaffected. The possible interactions of copper with prostaglandin-mediated inflammation and with free-radical-induced liver damage are discussed.     

Hepatic sinusoidal cell destruction in the development of intravascular coagulation in acute liver failure of rats

Rats received a dose of dimethylnitrosamine (DMN) or carbon tetrachloride (CCl4). In the liver of rats given DMN, apoptosis of fat-storing cells occurred at 7.5 h, and sinusoidal endothelial cell degeneration followed, with parenchymal cell necrosis after 9 h. Fibrin thrombi appeared in the sinusoids as well as in these necrotic areas after 12 h. In contrast, in the liver of rats given CCl4, parenchymal cell degeneration was seen after 6 h and necrosis with fibrin thrombi developed after 9 h. Fat-storing cells and endothelial cells were almost intact, and fibrin thrombi were not present in the sinusoids. SGPT values increased with decreased plasma levels of fibrinogen and antithrombin III and prolonged prothrombin time after 3 and 6 h, in the CCl4 and DMN models, respectively. An extensive reduction in plasma factor VIIIC levels and peripheral platelets was seen after 18 and 24 h, respectively, only in the DMN model. These results suggest that endothelial cells destruction can cause fibrin formation in the hepatic sinusoids in acute liver injury. Fat-storing cell injury may contribute to the destruction.     

Efficacy of a combined treatment with plasma exchange and cytostatics in macroglobulinemia

Monthly plasma exchange (PE) sessions have been carried out in 3 patients with advanced Waldenstr?m macroglobulinemia, in order to reduce electrophoretic M band under 2g/100 ml. When PE was combined to low doses of cytostatics (n = 18), 3 procedures per session were required to obtain a mean 57.4 +/- 12.3% IgM reduction, from 4.2 +/- 1.2 to 1.7 +/- 0.5 g/100 ml. A mean 61.5 +/- 13.1% IgM reduction, from 5.5 +/- 1.3 to 2.1 +/- 1 g/100 ml, was obtained in 64 procedures carried out as the only therapy in 12 sessions, with 5.3 procedures requirement per monthly session. IgM percent reduction 24 hours after PE was greater with combined treatment (45 +/- 9.7 vs. 28.9 +/- 15.4%; p = 0.001). The advantage of a combined treatment is therefore either a lowered PE requirement or a tapered maintenance cytostatic dosage.     

Tolerance of aged Fischer 344 rats against chlordecone-amplified carbon tetrachloride toxicity

We have investigated the effects of chlordecone 1(CD)+CCl4 combination in adult (3 months), middle aged (14 months), and old aged (24 months) male Fischer 344 (F344) rats. After a non-toxic dietary regimen of CD (10 ppm) or normal powdered diet for 15 days, rats received a single non-toxic dose of CCl4 (100 microl/kg, i.p., 1:4 in corn oil) or corn oil (500 microl/kg, i.p.) alone on day 16. Liver injury was assessed by plasma ALT, AST, and histopathology during a time course of 0-96 h. Liver tissue repair was measured by [3H-CH3]-thymidine (3H-T) incorporation into hepatic nuclear DNA and proliferating cell nuclear antigen (PCNA) immunohistochemistry. Hepatomicrosomal CYP2E1 protein, enzyme activity, and covalent binding of 14CCl4-derived radiolabel were measured in normal and CD fed rats. Exposure to CCl4 alone caused modest liver injury only in 14- and 24-month-old rats but neither progression of injury nor mortality. The CD+CCl4 combination led to 100% mortality in 3-month-old rats by 72 h, whereas none of the 14- and 24-month-old rats died. Both 3- and 14-month-old rats exposed to CD+Cl4 had identical liver injury up to 36 h indicating that bioactivation-mediated CCl4 injury was the same in the two age groups. Thereafter, liver injury escalated only in 3-month-old while it declined in 14-month-old rats. In 24-month-old rats initial liver injury at 6 h was similar to the 3- and 14-month-old rats and thereafter did not develop to the level of the other two age groups, recovering from injury by 96 h as in the 14-month-old rats. Neither hepatomicrosomal CYP2E1 protein nor the associated p-nitrophenol hydroxylase activity or covalent binding of 14CCl4-derived radiolabel to liver tissue differed between the age groups or diet regimens 2 h after the administration of 14CCl4. Compensatory liver tissue repair (3H-T, PCNA) was prompt and robust soon after CCl4 liver injury in the 14- and 24-month-old rats. In stark contrast, in the 3-month-old rats it failed allowing unabated progression of liver injury. These findings suggest that stimulation of early onset and robust liver tissue repair rescue the 14- and 24-month-old F344 rats from the lethal effect of the CD+CCl4 combination.     

Effect of luminal somatostatin on pentagastrin-stimulated gastric acid secretion in the rat

The perfused rat stomach was used to investigate the effect of intragastrically administered somatostatin (S-14) on basal and pentagastrin-stimulated gastric acid secretion. Intravenous injection of pentagastrin (10 micrograms/kg body wt) induced a peak acid output (PAO) of 6.3 +/- 0.45 mu eq H+. With luminal perfusion of the stomach by S-14 (100 micrograms X kg-1 X min-1), no significant inhibition of gastric acid secretion was observed (PAO: 5.9 +/- 0.6 mu eq H+). The same dose of S-14 administered intravenously significantly inhibited acid secretion (PAO: 1.7 +/- 0.4 mu eq H+) as did intravenous injection of neutralized S-14-containing gastric perfusate obtained by perfusion from a different rat stomach (PAO: 1.9 +/- 0.5 mu eq H+). Intravenous injection of a saline gastric perfusate containing no S-14 did not alter gastric acid secretion (PAO: 6.3 +/- 0.7 mu eq H+). It is concluded that S-14 does inhibit gastric acid secretion when administered systemically but not by intraluminal application.     

Interaction of hypoxia and carbon tetrachloride toxicity in hepatocyte monolayers

The toxicity of carbon tetrachloride (CCl4) in monolayer cultures of primary hepatocytes was investigated at oxygen concentrations that prevail in the liver under conditions that range from normoxia to hypoxia: 0.5, 1, 2, and 20% O2. CCl4 was administered in the vapor phase at concentrations that produce aqueous concentrations at 37 degrees C of 0.4, 2.0, and 4.0 mM. Damage was assayed by leakage of aspartate transaminase and the inclusion of Trypan Blue immediately after the 2-hr incubation and after an additional 6-hr incubation in 20% O2. Only in the case of 0.5% O2 and 4 mM CCl4 were the monolayers damaged (18%) immediately after the 2-hr exposure; all other exposed cells were undamaged at that time point and the dose response of cell death as a function of CCl4 and oxygen concentration was not evident until the 6-hr time point. The monolayers exposed to 4 mM CCl4 and 1, 2, or 20% O2 exhibited little immediate damage but were all 100% dead 6 hr later. The monolayers exposed to 2 mM CCl4 and 0.5, 1, 2, or 20% O2 were 53, 48, 40, and 22 +/- 2% dead after 6 hr, respectively. These results suggest that effects of CCl4 exposure, for example alterations in the function or synthesis of essential proteins, require several hours to affect cell viability.     

Comparison of 100 micrograms TRH intravenously and 40 mg TRH orally in euthyroidism (author's transl)

Euthyroid in-patients were investigated with 100 micrograms TRH given intravenously and with 40 mg TRH given orally in order to compare the sensitivity of intravenous and oral TRH-test. In all patients (age 15-91 years) maximal TSH-levels 180 min after 40 mg TRH are significantly (p less than 0.01) higher than maximal TSH-levels 30 min after 100 micrograms TRH. In patients under 50 years of age TSH-30 and TSH-180 are identical on average (6.42 +/- 0.29 microunits/ml vs. 6.53 +/- 0.54 microunits/ml/log x +/- SDlog). In patients older than 50 years TSH-30 is significantly lower (p less than 0.02) than in patients aging less than 50 years (2.11 +/- 0.88 microunits/ml vs. 6.42 +/- 0.29 microunits/ml). Both age-related groups demonstrate no significant difference (p greater than 0.2) in TSH-180 after 40 mg TRH (6.53 +/- 0.54 microunits/ml vs. 3.65 +/- 0.74 microunits/ml). 14 cases (34%) are low or non responders to 100 micrograms TRH intravenously, 8 (19.5%) only following 40 mg TRH orally. Patients with complete TSH-suppression (both iv. and oral TRH-test negative) show significant (p less than 0.05) higher ETR, T4 and fT4 than patients with partial TSH-suppression (iv. negative and oral TRH-test positive). The dose of 100 micrograms TRH does not increase the sensitivity of the test, the specificity is decreased in older patients, the dose of 100 micrograms TRH seems to be to low to yield reliable results. 40 mg TRH orally does not show any false positive findings, because TSH-stimulation is longer and more intensive. T3 increase three hours after orally TRH-application indicates an intact feedback mechanism.     

Isoenzymes in plasma of acute leukaemia patients: prevention of anthracycline cardiomyopathy

During the diagnosis and subsequent monitoring of patients with acute leukaemia, and of those with terminal blastic crisis in chronic myelogenous leukaemia, the activity of total LDH, ALAT, AP, ASAT as well as the isoenzymes LDH-H, LDH-M were measured in the plasma. Enzyme and isoenzyme activity of LDH, which differed in quantity at the various times of measurement, reflected the varying proliferation rate, depending upon the individual and upon the time of measurement and, consequently, the tumour cell mass.     

Endomyocardial biopsy of normal donor hearts before cardiac transplantation. A morphological and morphometrical study in 97 cases.

Endomyocardial biopsies from 97 normal donor hearts were examined. Morphometric analysis showed: mean myocyte diameter 22.21 +/- 6.93 mu, mean nuclear dimension 7.32 +/- 2.33 mu, mean nuclear/sarcoplasmic ratio 0.33 +/- 0.02. 31 biopsies showed enlarged myocytes (mean diameter 31.65 +/- 3.98 mu) with increased nuclear size (mean 10.45 +/- 1.39 mu), but preserved nuclear/sarcoplasmic ratio (mean 0.33 +/- 0.01). The mean age of these latter subjects was significantly higher. Endocardial thickness mean value was 17.73 +/- 4.58 mu, but in 28 cases the value exceeded the considered upper normal limit of 20 mu. Interstitial mononuclear cells were rare and randomly present. Interstitial fibrosis was observed in 15% and focal fibrosis in 27% of cases. Our results show that histology of biopsies from clinically normal hearts can widely vary, sometime overriding the pathologic boundaries. These apparently "benign" abnormalities should be kept in mind when specific pathologic substrates of cardiac diseases have to be defined.     

Cause and frequency of posttransfusion hepatitis after open-heart surgery

Summary A total of 1476 patients who underwent open-heart surgery between 1986 and 1988 participated in a prospective study examining posttransfusion hepatitis. They received a total of 8327 units of whole blood, packed erythrocytes, or fresh frozen plasma. The aminotransferase activities were measured preoperatively and 1, 2, 3, 4, 6, 9, 12, and 24 weeks after the operation. Thirty-four patients in all (2.3% of the transfused patients) developed posttransfusion hepatitis, which could be identified as hepatitis B in 1 patient and hepatitis C in 14 patients. No cause for posttransfusion hepatitis could be found in 19 cases (hepatitis of unknown origin). Hepatitis C became chronic in 5 patients. In contrast to hepatitis C, the 19 patients with hepatitis of unknown origin all showed a milder clinical course with lower maximal aminotransferase activities and a shorter duration of the hepatitis. A chronic course was not observed among them. The cause of hepatitis of unknown origin is discussed.Abbreviations Hep of UO hepatitis of unknown origin - ELISA enzyme-linked immunosorbent assay - ALAT alanine aminotransferase - ASAT asparagine aminotransferase - AT aminotransferase - AT-A aminotransferase activity - CMU cytomegalovirus - EBU Epstein-Barr virus     

Central control of peripheral circulating somatostatin in dogs: effect of 2-deoxyglucose

Circulating plasma somatostatin concentrations are known to fluctuate in response to nutrients and hormones. However, little is known about neural or central nervous system (CNS) control of somatostatin secretion. To test whether peripheral circulating somatostatin is influenced by a central stimulus, 2-deoxyglucose (37.5 mg/kg) was infused into a lateral cerebral ventricle of six conscious dogs over a period of 15 min. Plasma somatostatin levels rose from a base line of 105 +/- 6 pg/ml (mean +/- SE) to a peak of 154 +/- 10 pg/ml (P less than 0.005) at 30 min after the onset of the infusion. Somatostatin levels were still significantly elevated (P less than 0.025) at 60 min (119 +/- 6 pg/ml) and thereafter gradually returned toward base line. Plasma glucose and glucagon levels increased in response to intraventricular 2-deoxyglucose. Glucose concentrations rose from 105 +/- 5 mg/dl to peak at 203 +/- 16 mg/dl (P less than 0.005) at 80 min and remained elevated to 120 min. The concentration of plasma glucagon increased from 41 +/- 6 to 92 +/- 18 pg/ml at 60 min (P less than 0.05) and then declined. In marked contrast to intraventricular 2-deoxyglucose, similar concentrations of 2-deoxyglucose administered intravenously (n = 4) resulted in a slight fall in plasma somatostatin. Intraventricular saline did not result in a change in plasma somatostatin. It is concluded that peripheral circulating somatostatin may be susceptible to central nervous system control.     

永生化人肝细胞治疗小鼠急性肝衰竭的实验研究

目的评价微载体培养的永生化人肝细胞(immortalized human hepatocytes,IHH)对裸鼠急性肝衰竭(acute liver failure,ALF)的治疗作用。方法采用腹腔注射四氯化碳(CCl4)建立裸鼠ALF模型,将微载体培养的IHH注射于裸鼠腹腔内进行腹膜透析治疗,并设未治疗组和空微载体治疗组,比较各组动物的肝功能损伤指标,肝组织病理和动物生存率。结果腹腔注射CCl4后24h,所有动物的转氨酶升高达正常值10倍以上,未治疗组和空微载体治疗组动物48h内全部死亡,肝组织检查发现肝细胞广泛坏死;而采用IHH微载体治疗组,48h的动物生存率为83.3%,持续14d后转为长期存活。存活的动物转氨酶降至正常水平,肝组织检查显示肝细胞坏死显著减少,免疫组织化学染色显示腹腔微载体上肝细胞仍存活,并表达人肝细胞特异蛋白α1-抗胰蛋白酶(α1-antitrypsin,α1-AT)。结论采用微载体培养的IHH腹腔透析治疗小鼠ALF是有效的,表明IHH适合作为生物人工肝的细胞材料。     

Changes in several plasma and urinary components in marathon runners

33 blood and urinary components were titrated in 11 men 5 days before a marathon race (42 km), just before the start of the race, 1/4 hour after the arrival, the following day and 5 days after the race. On arrival, or/and the following day, even still 5 days later, we observed an increase of: natremia, kaliemia, blood proteins, hematocrit, aldosteronemia, plasma renin activity, uricemia, creatininemia, blood cortisol, myoglobinemia, blood lactic acid, total enzymatic CK activity, enzymatic ASAT activity, urinary elimination of creatinin, urea and 3 methylhistidine. On the contrary, a decrease of plasma total CO2 and blood testosterone levels were observed. These biochemical modifications are the consequence of hydro-mineral losses, muscular necroses, alteration of energetic metabolism with increase of the protein catabolism.     

Pharmacokinetics of doxycycline in broiler chickens

Doxycycline was given to two groups of eight chickens at a dose of 20 mg/kg of body weight, intravenously (i.v.) or orally. Plasma concentration was monitored serially for 12 h after each administration. Another group of 30 chickens was given 20 mg/kg orally every 24 h for 4 days, and plasma and tissue concentrations determined serially after the last administration. Concentrations of doxycycline were measured using high-performance liquid chromatography. Pharmacokinetic variables were calculated, using a two-compartment open model. The elimination half-life and the mean residence time for plasma were 6.03 +/- 0.45 and 7.48 +/- 0.38 h, respectively, after oral administration and 4.75 +/-0.21 and 2.87 +/-0.11 h, respectively, after i.v. administration. After single oral administration, doxycycline was absorbed rapidly, with T(max) of 0.35 +/- 0.02 h. Maximum plasma concentration was 54.58 +/- 2.44 mu/ml. Oral bioavailability of doxycycline was found to be 41.33 +/- 2.02%. Doxycycline was widely distributed in tissues and considerable concentrations were found following oral administration of 20 mg/kg on four successive days. The results indicate that doxycycline concentrations were cleared slowly and were at or below the accepted drug tolerance levels in the marker tissues within 5 days after dosing.