Although heparins are usually injected intravenously or subcutaneously, antithrombotic activity is observed in rat models following single oral heparin doses. Since repetitive dosing is usually needed for thromboprophylaxis, study objectives were to determine whether repetitive oral heparin prevented arterial thrombosis and to compare effectiveness to subcutaneous administration. Wistar rats were given subcutaneous or oral unfractionated heparin ([UFH] 1 mg/kg per 48 h), low-molecular-weight heparin ([LMWH] tinzaparin, 0.1 mg/kg per 12 h), or saline for 30 days. On the last day, thrombosis was initiated by placing 30% FeCl(3)-soaked filter paper on the distal carotid. Subsequent flow measurements, for a 60-minute period, included recorded time of initial thrombus formation (time till thrombus begins [TTB]), and time until carotid occlusion (time till occlusion [TTO]). The formed thrombus was dried and weighed. The activated partial thromboplastin time (aPTT), anti-factor Xa, and antithrombin activity were determined from the plasma. Both oral and subcutaneous heparins significantly increased TTB and TTO. Time of initial thrombus formations were 12.6 ± 1.1, 21.2 ± 2.2, 25.3 ± 3.9, 21.7 ± 3.1, and 21.3 ± 1.7 minutes and TTOs were 29.3 ± 3.6, 54.8 ± 4.0, 60.0 ± 0.3, 56.7 ± 3.3, and 58.3 ± 1.7 minutes (mean ± SEM) for control, subcutaneous UFH, oral UFH, subcutaneous LMWH, and oral LMWH, respectively. Thrombus weight was 2.52 ± 0.29 g in control and was reduced to 43%, 23%, 33%, and 28% of control weight for subcutaneous UFH, oral UFH, subcutaneous LMWH, and oral LMWH, respectively. Thrombus weight was significantly less for oral compared to subcutaneous UFH. The aPTT for oral UFH, and anti-factor Xa activity in the LMWH-treated groups were significantly greater than control (two-tailed t tests). These findings confirm that orally administered heparins are absorbed. Repeated treatment with oral heparin showed similar antithrombotic activity compared to subcutaneous heparin. Oral heparin use for arterial thromboprophylaxis should be further investigated.

BACKGROUND AND PURPOSE:An oral, single dose of 7.5 mg kg(-1) of unfractionated heparin (UFH) reduces thrombosis by 50% in a rat model of venous thrombosis. As long-term use is required clinically, our objectives were to study the antithrombotic effects following repeated oral UFH administration. EXPERIMENTAL APPROACH:Bovine lung UFH was administered by oral gavage to rats in 3 doses of 7.5 mg kg(-1) each 12, 24, 48, and 72 h apart; and in 3 or 15 doses of 1 mg kg(-1) every 48 h. The last dose was given immediately after thrombus initiation where 10% formalin in methanol was applied to the jugular vein. The vessel was examined for thrombosis 4 h later. Amounts of heparin in tissue and endothelium, and plasma anticoagulant activity were measured. KEY RESULTS:When 3 x 7.5 mg kg(-1) heparin was given, thrombotic incidence was most reduced at 48 h dose-intervals and was significantly less than single dose treatment. There was a negative correlation between endothelial heparin content and thrombotic incidence, but not anticoagulant activity. When 3 doses of 1 mg kg(-1) every 48 h were given, thrombotic incidence was similar to single dose treatment. When 15 doses were given, total thrombotic incidence was less than for 3 doses and was similar to that after s.c. administration. CONCLUSIONS AND IMPLICATIONS:Antithrombotic activity increased with repeated doses of oral UFH, with antithrombotic effects similar to s.c. administration. Antithrombotic activity was related to heparin on endothelium.

On the basis of suggested clinical efficacy in an uncontrolled study in ninety-seven patients with unstable angina, an animal study was conducted to investigate antithrombotic synergy between orally administered heparin and arginine. A rat venous thrombosis model tested the difference in thrombus formation when heparin (7.5 mg/kg) and arginine (113 mg/kg) were administered, alone or in combination, by stomach tube with a minimum of 20 rats/group. Oral heparin, arginine, and heparin plus arginine reduced thrombus formation by 50%, 75%, and 90%, respectively,when compared to saline administration. Heparin was recovered from endothelium, yet there was little or no observable plasma anticoagulant activity. An orally administered low-molecular-weight anticoagulant glycosaminoglycan mixture, sulodexide (7.5 mg/kg), showed an 88% reduction in stable thrombus formation when administered alone but showed no synergy with oral arginine. A 28-day study with oral sulodexide (2.9 mg/kg) and arginine (43.9 mg/kg), 20 rats/group, showed antithrombotic activity with minimal anticoagulant activity indicating suitability for long term treatment. These findings suggest the endothelial localization of heparin and a synergistic antithrombotic effect for orally administered heparin and arginine.

Antithrombotic activity and heparin with endothelium are observed in rats when heparin is administered by the oral route. Peak endothelial concentrations at 6 min suggest rapid absorption. To identify the site of absorption, stomach and duodenum were isolated by tying the pyloric sphincter of male Wistar rats and heparin (unfractionated bovine lung, 60 mg/kg) was administered by stomach tube or injected into the duodenum. Heparin in plasma and aortic endothelium, collected within 15 min, was determined by densitometry following agarose gel electrophoresis with toluidine blue staining. Heparin was recovered in 5 of 10 endothelial (0.136+/-0.068 microg/cm(2)) and 6 of 10 plasma (0.06+/-0.02 microg/ml) samples when administered in the stomach and in 0 of 9 endothelial and 2 of 9 plasma (0.02+/-0.02 microg/ml) samples when injected into the duodenum. To further study heparin distribution, stomach layers were separated and analysed 15 min and 4h following heparin administration by stomach tube. Heparin was recovered in muscle and mucosal layers as well as washes indicating that heparin passes through stomach tissue. Heparin was also recovered from the portal vein, endothelium and lung. These results indicate that heparin is absorbed following oral administration and that the stomach is an important site of absorption.

Although heparin is not generally administered orally, the results of studies involving rats suggest that heparin is absorbed, with low levels in plasma but extensive distribution to the endothelium. To determine whether evidence of absorption after oral administration can also be demonstrated in human subjects, we administered unfractionated porcine heparin in a single dose of 1000 U/kg to 6 healthy human subjects. Plasma anticoagulant activity was monitored between 5 minutes and 72 hours after administration, and chemical heparin concentrations were determined in 24-hour urine samples for as long as 120 hours after administration. Plasma anticoagulant activity, determined by anti-Xa activity, increased as soon as 5 minutes after heparin administration, peaked at 120 minutes, and was still increased 72 hours after administration. Values were significantly greater 15 minutes to 48 hours after administration compared with values before administration (paired t test). Mean activated partial thromboplastin time and Heptest values in subjects given heparin were greater than those in controls 15 and 30 minutes and 5 to 120 minutes after administration, respectively. Heparin was recovered from urine as long as 120 hours after administration (the latest time point at which samples were collected); greater amounts were excreted between 48 and 120 hours than before 48 hours. Recovery from both plasma and urine suggest that unfractionated heparin administered orally is absorbed in human subjects, is widely distributed, and is found in the body at least 120 hours after administration. Because heparin is readily bound to endothelium, recovery from plasma and urine likely reflect considerable amounts with endothelium, as has been observed in other species.

Heparins are antithrombotic drugs given by intravenous and subcutaneous routes. However, we have observed that heparins have antithrombotic activity in a rat model when administered orally despite low plasma levels, with low molecular weight heparins (LMWHs) being effective at lower single doses than unfractionated heparins (UFH). Since LMWHs may have other pharmaceutical uses and little is known regarding the pharmacokinetics of oral LMWHs, our objectives were to determine the distribution of the LMWH tinzaparin (Logiparin) following oral dosing. To study distribution at different doses, 0.025-15 mg/kg tinzaparin was given by stomach tube to rats. Gut and non-gut tissues were sampled 4 h later. In a time course study, plasma and tissue samples were collected at eight time points within 24 h after oral administration (60 mg/kg, 4 rats/time interval). Accumulated urine and faeces were collected over 4 and 24 h using metabolic cages. Gut tissue and washes, faeces, urine and non-gut tissue were extracted and analysed for heparin by agarose gel electrophoresis with toluidine blue staining. Activated partial thromboplastin time (APTT) and anti-Xa activity, by Heptest and chromogenic assay, estimated plasma tinzaparin concentrations. Stomach and lung tinzaparin concentrations demonstrated a dose-effect. Peak concentrations in tissue and washes of stomach, duodenum, jejunum, ileum and colon were at 6-30, 15-30, 30 min, 2 and 4 h, respectively. Amounts found at peak times in combined tissue and washes accounted for 46% and 0.5% in stomach (15 min) and colon (4 h), respectively. Tinzaparin was recovered from liver, lung, endothelial samples, and urine at 24 h, but not in faeces. Non-significant increases were seen in APTT and the Heptest, however, anti-Xa activity was significantly greater than control at all times examined, peaking at 2 h. No bleeding was observed. Results are consistent with oral absorption of tinzaparin with wide tissue distribution, likely on endothelium with little in plasma, as previously observed for UFH. Oral administration of LMWHs should be further studied.

Summary Our previous studies demonstrated that orally administered heparins prevent thrombosis in a rat jugular vein thrombosis model, where bovine unfractionated heparin (UFH) and the low molecular weight heparin tinzaparin reduced thrombotic incidence by 50% at 7.5 and 0.1 mg/kg, respectively. Our objectives were to determine if similar antithrombotic effects of oral heparin could be observed in an arterial thrombosis model. In this model, filter paper soaked in 30% ferric chloride was applied to the exposed rat carotid artery. A flowmeter recorded blood flow over a 60 min period determining time when the thrombus began forming (TTB) and time till occlusion (TTO). Immediately following, the thrombus was removed, dried and weighed 24 h later. Bovine UFH (7.5 mg/kg), tinzaparin (0.1 mg/kg) or saline was administered by stomach tube at 2, 5 and 25 h prior to thrombus initiation. TTB was significantly increased when UFH was given at 5 and 25 h but not 2 h prior, and when tinzaparin was given at 5 but not 2 or 25 h prior compared to rats given oral saline. TTO was significantly increased for both UFH and tinzaparin when given 5 and 25 h but not 2 h prior (one-way ANOVA). There was no difference in TTO and TTB between UFH and tinzaparin treated groups. A trend in reduction in thrombus weight was observed for UFH at 5 and 25 h prior and tinzaparin at 5 h prior to thrombus initiation (one-way ANOVA). Although no significant changes were observed in activated partial thromboplastin times, Heptest or anti-Xa activity from plasma of heparin treated rats, endothelial heparin concentrations were significantly greater than controls for UFH at 5 h and for tinzaparin at 2, 5, and 24 h. Thus, heparins administered by the oral route are effective antithrombotic agents in arterial as well as venous models.

Purpose. Aluminum sucrose octasulfate (SOS) is used clinically to prevent ulcers. Under physiologic conditions, the sodium salt of this drug can be formed. Our objective was to determine whether sodium SOS was absorbed when administered orally. In addition to furthering our understanding of aluminum SOS, this study also aimed to clarify how other polyanionic drugs, such as heparin and low-molecular-weight heparins, are absorbed.

Previous studies in rats demonstrated that orally administered, unfractionated bovine lung heparin is absorbed and has a dose-dependent antithrombotic effect. The objective of this study was to determine if an oral low molecular weight heparin had a similar antithrombotic effect in the same model. Thrombosis was induced in rats by application of 10% formalin in 65% methanol to the exposed jugular vein. Immediately following, saline, unfractionated heparin (3.3–60 mg/kg) or the low molecular weight heparin, Logiparin (0.025–15 mg/kg; 20–30 rats per group) was placed in the stomach and 4 h later the jugular vein was inspected for a thrombus. Compared to saline, oral Logiparin reduced the incidence of thrombosis at all doses with a dose-dependent effect suggested. A significant increase was observed in the activated partial thromboplastin time and in plasma heparin concentrations, determined by AccuclotTM Heptest® and anti-factor Xa chromogenic assay for rats given oral Logiparin versus saline. A dose-dependent increase in plasma heparin concentration was observed when estimated by the anti-Xa chromogenic assay. Heparin was recovered in 9% of aortic endothelial samples when ≧0.8 mg/kg Logiparin was administered. A 50% reduction in thrombosis was observed at 0.1 mg/kg for oral Logiparin versus 7.5 mg/kg for unfractionated bovine lung heparin indicating that oral Logiparin is an effective antithrombotic agent at doses lower than unfractionated heparin. Orally administered low molecular weight heparin may be useful for the prevention and treatment of thrombosis.

Distribution and antithrombotic activity of orally administered unfractionated porcine heparin were studied. [14C]Heparin was prepared by de-N-acetylation of porcine mucosal heparin followed by re-N-acetylation, using [14C]acetic anhydride. [14C]Heparin and (or) cold heparin (60 mg/kg) were administered by stomach tube to male Wistar rats. Blood, all levels of gut and gut contents, liver, lung, spleen, kidney, and aortic and vena caval endothelium were collected under deep anesthesia at 3, 6, 15, 30, and 60 min and 4 and 24 h (6 rats/group) after administration. Urine and feces were collected at 24 h, using metabolic cages. In three additional rats, drugs were administered in gelatin capsules. Tissues listed above and tongue, esophagus, trachea, brain, heart, thymus, bile ducts, vena caval and aortic walls, ureters, bladder, samples of muscle, skin, hair, and bone marrow were collected at 24 h. Radioactivity and chemical heparin, measured by agarose gel electrophoresis, were observed in all tissues examined as well as gut washes, plasma, urine, and feces. Radiolabel recovered was confirmed to be heparin by autoradiograms of gradient polyacrylamide electrophoretic gels. [14C]Heparin and chemical heparin in gut tissue suggest a transit time of 4 h. Porcine or bovine heparin (7.5 mg/kg), administered by stomach tube, decreased the incidence of thrombosis induced by applying 10% formalin in 65% methanol to the exposed jugular vein of rats. Heparin isolation from non-gut tissue, endothelium, urine, and plasma and the observed antithrombotic effect are consistent with oral bioavailability.

Preliminary in vivo studies suggested that oral dextran sulfate was poorly absorbed, but investigations were limited by inadequate methods for measuring the drug in the body. To determine absorption in HIV-positive subjects, hydrogenated dextran sulfate, average molecular weight 8000 (Usherdex 8), was orally administered in a short-term (single dose, 4 g/day for 5 days, 7 subjects) and in a long-term study (1 g, 4 times per day for 29 to 335 days, 8 subjects), which was a continuation of the short-term study with the inclusion of an additional subject. When an agarose gel electrophoresis technique with toluidine blue staining was used, the drug was recovered from plasma (67%, peak 2.2 μg/mL) and circulating peripheral blood lymphocyte (PBL) samples (50%, peak 333 μg/L blood) obtained at 5 and 15 minutes and 1, 3, 6, and 24 hours after the first day's dose and from plasma (56%) and PBL samples (38%) obtained 5 minutes after administration on 4 subsequent days in the short-term study. In the long-term study, the drug was found in plasma (67%, peak 2.4 μg/mL) and PBL samples (25%, peak 126 μg/L blood) obtained at monthly visits within 4 hours of the last dose. The drug was found in all urine samples from all subjects in both studies (short-term study, 24-hour samples up to 4 days after the final dose; long-term study, monthly samples within 4 hours of the last dose). In the long-term study, bone marrow preparations from 3 subjects showed metachromatic inclusions present in reticular cells when the cells were stained with toluidine blue, indicating the presence of sulfated polyanions. A significant rise in activated partial thromboplastin time and a drop in platelet count (P <.025) were demonstrated, with thrombocytopenia developing in 3 patients. Mild-to-moderate gastrointestinal disturbances were experienced by 6 subjects in the short-term study and by all subjects in the long-term study. One subject experienced mild central nervous system symptoms in the short-term study. These results indicate that dextran sulfate is absorbed after oral administration; therefore, further studies on its efficacy, particularly in the early stages of the disease, along with additional observations on its toxicity, are warranted. (J Lab Clin Med 1999;133:161-70)

Although heparin is believed to be poorly absorbed orally, we recently demonstrated that oral heparin rapidly enters the circulation, with most of the drug being taken up by endothelium. To determine the effective antithrombotic dose of oral heparin, we induced thrombosis by applying 10% formalin in 65% methanol to exposed rat jugular vein. Saline or heparin, at doses ranging from 3.25 to 60 mg/kg, was immediately placed in the stomach; 4 h later, the vein was inspected for a thrombus. A dose-dependent decrease in thrombosis was observed with oral heparin. Although there was little change in anticoagulant activity as measured by the activated partial thromboplastin time (APTT) of plasma samples taken 4 h after administration, a significant dose effect was demonstrated by regression analysis. Heparin could be demonstrated chemically in 52% of plasma samples and in 38% of aortic or vena caval endothelial samples. A significant dose effect was observed in aortic endothelial heparin concentrations, with amounts 1,000-fold that determined in plasma. These results indicate that oral heparin exhibits antithrombotic activity in a dose-dependent manner, with low levels in plasma.