Partial Thromboplastin Time Test Research Articles

Anticoagulant Activity of Rhamnan Sulfate Isolated from Commercially Cultured <i>Monostroma nitidum</i>

The green seaweed, <i>Monostroma nitidum</i>, is widespread in Japan. In Okinawa Prefecture, the production of seaweed is performed using culture-nets that are seeded artificially. Algae contain a soluble polysaccharide, rhamnan sulfate. To estimate its applicability, the anticoagulant activity of rhamnan sulfate was investigated. Rhamanan sulfate was fractionated by ion-exchange chromatography on a DEAE-sepharose column, and two fractions (A and B) were obtained. Partially hydrolyzed rhamnan sulfates with different molecular mass (C1, C2 and C3) were also prepared. The activated partial thromboplastin time (APTT) test, prothrombin time (PT) and thrombin time (TT) were applied using human plasma and compared with standard heparin (174 units/mg). The native rhamnan sulfate (molecular mass, 630 kDa; sulfuric acid content, 22.7%), fraction A (12.4%) and fraction B (27.8%) showed approximately 73% APTT activity in comparison with that of standard heparin, but fraction C2 (molecular mass, 450 kDa) had a higher activity than that of the standard (107%). On the other hand, in the PT assay, all fractions except fraction C2 and C3 (370 kDa) showed higher activity approximately 120-155% greater than that of standard heparin. The TT activity of rhamnan sulfate depended on the sulfate content, and that of fraction B, which has high sulfuric acid content (27.8%), was 135-173% greater than that of heparin. The sulfate groups of L-rhamnosyl residues and carboxyl group of D-glucuronosyl residue on the trisaccharide side chains of the rhamnan sulfate might interact strongly with the active site of thrombin molecules. The results and discussion suggested that rhamnan sulfate from commercially cultured <i>Monostroma nitidum</i> could be a potential anticoagulant polysaccharide.

In vitro and in vivo anti-coagulant activity and toxicological studies of marine sulfated glycosaminoglycans

The present study aimed to characterize and evaluate the in vitro and in vivo anticoagulant activity of sulfated glycosaminoglycans from the skins of smooth hound (SHSG) and grey triggerfish (GTSG). The analysis of SHSG and GTSG with acetate cellulose electrophoresis in Zn-acetate revealed the presence of hyaluronic acid (HA), chondroitin sulfate (CS) and dermatan sulfate (DS). Both glycosaminoglycans were evaluated for their in vitro anticoagulant activities using activated partial thromboplastin time (aPTT), thrombin time (TT) and prothrombine time (PT) tests. SHSG and GTSG and calciparin were tested as in vivo anticoagulants by subcutaneous (s.c) injection to adult female Wistar rats in a concentration of 75mg/kg of body weight. The administration of SHSG, GTSG and calciparin to rats induced a significant decrease of platelet rates compared to the control. The aPTT assay of SHSG and GTSG was prolonged 1.3 and 1.23-fold respectively compared with the control. Toxicity studies were performed to investigate whether or not SHSG and GTSG can cause pathological changes in the liver, proteins and DNA. The concentration and catalytic activity of liver oxidative stress markers and enzymes, respectively, as well as the observed hepatic morphological changes indicated that calciparin induced hepatic toxicity and oxidative damage in the liver. The higher activity and lower toxicity of SHSG and GTSG recommended these compounds as a better drug candidate than calciparin.

The confusion continues: results from an American Association for the Surgery of Trauma survey on massive transfusion practices among United States trauma centers.

Massive transfusion practices have undergone several recent developments. We sought to examine institutional practices guiding hemostatic resuscitation in the setting of massive hemorrhage. A 37-question online survey was sent to American Association for the Surgery of Trauma members. A total of 191 surgeons from 125 institutions completed the survey. Level I and II centers composed 70 and 18% of responding sites, respectively. A total of 123 institutions have a massive transfusion protocol (MTP); 54% report having an MTP for less than 5 years. The number of coolers and units of red blood cells, plasma, and platelets are highly variable. Tranexamic acid is part of the MTP at 64% of centers; 26% continue to use recombinant activated Factor VII. MTP activation occurs more than five times per month at 32% of centers. MTPs are utilized for nontrauma patients in 82% of institutions. Point-of-care prothrombin time, international normalized ratio, and partial thromboplastin time testing is utilized in 37% of institutions. Only 9% routinely utilize thromboelastography or rotational thromboelastometry (TEG/ROTEM) within their MTP. Just 7% use a validated scoring system to guide MTP activation. The incorporation of TEG/ROTEM into the MTP is associated with the use of a scoring system in regression analysis (p=0.024). Most institutions regularly activate recently implemented MTPs for trauma and nontrauma indications; however, few use validated scoring systems for MTP activation. MTP content is highly variable. Few institutions use TEG, while most have incorporated tranexamic acid into their protocol. The lack of consistent practices underscores the need for outcome-based studies to guide transfusion practices.

&lt;b&gt;Extraction and structural properties of &lt;i&gt;Acanthophora muscoides &lt;/i&gt;(Rhodophyceae) extracellular matrix sulfated polysaccharides and their effects on coagulation

Acanthophora muscoides (Rhodophyta) contains structurally heterogeneous sulfated polysaccharides (Am-SPs) with pharmacological importance; however, its matrix SPs composition has not been still extensively investigated. This study sequentially extracted and compared the structural features and the in vitro anticoagulant effects of the Am-SPs. Papain-extraction sequence yielded Am.E-1, Am.E-2 and Am.E-3 containing differences among the relative proportions of sulfate (26.18-33%) and hexoses (42.02-60.67%) based on chemical analyses. One- ( 1 H) and two-dimensions ( 1 H/ 13 C) nuclear magnetic resonance experiments showed very complex Am-SPs composed of alternating 4-linked-α-galactopyranosyl units and 3-linked-β-galactopyranosyl units presenting variable sulfation, CH 3 substitutions and3,6-anhydro-α-L-galactose units and pyruvated-D-galactose residues, respectively, typical of agarocolloids. Different chromatographic profiles (DEAE-cellulose) were observed, with fractions (Am I, Am II and Am III eluted with 0.5, 0.75 and/or 1 M of NaCl, respectively) revealing charge density patterns and distinct mobility to heparin by agarose-electrophoresis and, when analyzed by polyacrylamide-electrophoresis, a dispersive migration and similar mobility as chondroitin-6-sulfate for Am I fractions were noted. Regarding the activated partial thromboplastin time test, fractions had no virtually anticoagulation (1.47→3.07 IU mg -1 ) in comparison with 193 IU mg -1 heparin. Therefore, Am-SPs show significantly lower anticoagulation than heparin.

A vascular tissue engineering scaffold with core–shell structured nano-fibers formed by coaxial electrospinning and its biocompatibility evaluation

In this article, a tubular vascular tissue engineering scaffold with core–shell structured fibers was produced by coaxial electrospinning at an appropriate flow rate ratio between the inner and outer solution. PCL was selected as the core to provide the mechanical property and integrity to the scaffold while collagen was used as the shell to improve the attachment and proliferation of vascular cells due to its excellent biocompatibility. The fine core–shell structured fibers were demonstrated by scanning electron microscope and transmission electron microscope observations. Subsequently, the collagen shell was crosslinked by genipin and further bound with heparin. The crosslinking process was confirmed by the increasing of tensile strength, swelling ratio and thermogravimetric analysis measurements while the surface heparin content was characterized by means of a UV-spectrophotometer and activated partial thromboplastin time tests. Furthermore, the mechanical properties such as stitch strength and bursting pressure of the as-prepared scaffold were measured. Moreover, the biocompatibility of the scaffold was evaluated by cytotoxicity investigation with L929 cells via MTT assay. Endothelial cell adhesion assessments were conducted to reveal the possibility of the formation of an endothelial cell layer on the scaffold surface, while the ability of smooth muscle cell penetration into the scaffold wall was also assessed by confocal laser scanning microscopy. The as-prepared core–shell structured scaffold showed promising potential for use in vascular tissue engineering.

Effect of Subcutaneous Unfractionated Heparin Prophylaxis on Activated Partial Thromboplastin Time: A Retrospective Evaluation

Study ObjectiveCharacterize the incidence of elevated aPTT results in patients treated with prophylactic, subcutaneous unfractionated heparin (UFH). DesignRetrospective, cohort analysis. SettingSingle-center, university hospital. MeasurementsEvaluation of 257 patients with activated partial thromboplastin time (aPTT) testing both prior to and following subcutaneous (SC) unfractionated heparin (UFH) therapy. Main ResultsEvaluated patients received UFH 5000 units every 8 hours. Baseline aPTT values were within the normal range (mean±SD, 32.0±8.5 seconds). After initiation of UFH, aPTT values increased (mean±SD, 37.6±15.2 seconds). After 24 hours of SC UFH, mean aPTT values (mean±SD, 38.6±15.5) exceeded the normal laboratory range (23.3–35.7 seconds). An elevated aPTT result after UFH was associated with baseline aPTT, length of therapy, and weight-based UFH dose. A significant association was not identified between aPTT elevation and age, race, sex, history of liver disease, type of admission, or transfusion of blood products. ConclusionsTreatment with UFH resulted in a small, but significant, increase in aPTT.

IN VITRO HAEMOSTATIC ACTIVITY OF RHODOMYRTUS TOMENTOSA (AITON) HASSK. AQUEOUS LEAF EXTRACT

Potential of Rhodomyrtus tomentosa as a native anticoagulant been tested by evaluating the possible haemostatic effect of Rhodomyrtus tomentosa aqueous leaf extract in vitro. In vitro coagulation assays such as Thrombin time (TT), Prothrombin time (PT) and Activated partial thromboplastin time (APTT) tests were performed on normal coagulation control plasma spiked with different concentrations of the leaf extract (10 – 100%). Total tannin and flavonoid for all different concentrations of leaf extract was measured using spectrophotometer. The results were analyzed using one way ANOVA followed by post-hoc Dunnet’s and Pearson’s correlation. The results showed that TT, PT and APTT of normal coagulation control plasma spiked with extract were markedly prolonged. A significant anticoagulant (p &lt; 0.05) showed by PT was at lowest concentration (10%), TT was at concentration 50 – 100% and APTT was at concentration 80% - 100%. The clotting times for TT, PT and APTT were significantly correlated (p &lt; 0.05) with total tannin and flavonoid. In summary, a potent anticoagulant effect of Rhodomyrtus tomentosa significantly higher in extrinsic pathway (PT) and the bioactive compound of tannin and flavonoid attributes to the anticoagulant activity of Rhodomyrtus tomentosa.

Anticoagulant properties and cytotoxic effect against HCT116 human colon cell line of sulfated glycosaminoglycans isolated from the Norway lobster (Nephrops norvegicus) shell.

Sulfated glycosaminoglycans (SGNL) were extracted for the first time from Norway lobster (Nephrops norvegicus) shell. The monosaccharide composition analysed by GC/MS revealed the presence of galacturonic acid, glucuronic acid, N-acetylgalactosamine and N-acetylglucosamine. The analysis of SGNL with acetate cellulose electrophoresis in Zn-acetate revealed the presence of heparan sulfate (HS) and dermatan sulfate (DS). SGNL were evaluated for their anticoagulant activities using activated partial thromboplastin time (aPTT), thrombin time (TT) and prothrombine time (PT) tests. After 21h incubation, HCT116 cell proliferation was inhibited (p<0.05) between 39.7 and 54.8% at 1.5-7.5mg/mL of SGNL. SGNL don't show hemolytic activity towards bovine erythrocytes and no cytotoxicity against the normal lymphocytes. The antiproliferative efficacy of these lobster glycosaminoglycans were probably related with the higher sulfate content. SGNL demonstrated promising antiproliferative and anticoagulant potential, which may be used as a novel, effective and promising antithrombotic agent.

Establishing the heparin therapeutic range using aPTT and anti-Xa measurements for monitoring unfractionated heparin therapy.

BackgroundUnfractionated heparin (UFH) has unstable pharmacokinetics and requires close monitoring. The activated partial thromboplastin time (aPTT) test has been used to monitor UFH therapy for decades in Korea, but its results can be affected by numerous variables. We established an aPTT heparin therapeutic range (HTR) corresponding to therapeutic anti-Xa levels for continuous intravenous UFH administration, and used appropriate monitoring to determine if an adequate dose of UFH was applied.MethodsA total of 134 ex vivo samples were obtained from 71 patients with a variety of thromboembolisms. All patients received intravenous UFH therapy and were enrolled from June to September 2015 at Gyeongsang National University Hospital. All laboratory protocols were in accordance with the Clinical and Laboratory Standards Institute guidelines and the College of American Pathologist requirements for aPTT HTR.ResultsAn aPTT range of 87.1 sec to 128.7 sec corresponded to anti-Xa levels of 0.3 IU/mL to 0.7 IU/mL for HTR under our laboratory conditions. Based on their anti-Xa levels, blood specimen distribution were as follows: less than 0.3 IU/mL, 65.7%; 0.3–0.7 IU/mL (therapeutic range), 33.6%; and more than 0.7 IU/mL, 0.7%. No evidence of recurring thromboembolism was observed.ConclusionUsing the conventional aPTT target range may lead to inappropriate dosing of UFH. Transitioning from the aPTT test to the anti-Xa assay is required to avoid the laborious validation of the aPTT HTR test, even though the anti-Xa assay is more expensive.

Effect of solvents extraction on HPLC profile of phenolic compounds, antioxidant and anticoagulant properties of Origanum vulgare

Origanum vulgare is traditionally used as food and medicinal plant. In the present study, the influence of three solvents; methanol, acetone and dichloromethane on total phenolic content, total flavonoid content, HPLC profile of phenolic compounds, antioxidant, and anticoagulant activities of O. vulgare were evaluated. Methanol extract showed the highest total phenolic content among the other two extracts; however the highest total flavonoid content was detected in acetone extract. HPLC results revealed the presence of eleven phenolic compounds and the methanol extract had the highest extraction capacity from quercetin, rutin and catechin as compared to acetone and dichloromethane extracts. All extracts exhibited both antioxidant and anticoagulant activities and methanol extract was superior compared to other extracts in the antioxidant activity while the acetone and dichloromethane extracts exhibited the highest anticoagulant activity in prothrombin time (PT) and activated partial thromboplastin time (APTT) tests. This study showed that extracts of O. vulgare may be considered as an important source for natural antioxidant and anticoagulant agents for application in pharmaceutical and food industries.

The Effect of Time and Temperature Variables on Some Routine Coagulation Tests among Subjects of African Descent in Sokoto, North Western Nigeria

This study evaluated the effects of time and temperature variables on routine Pro-thrombin Time test and Activated Partial Thromboplastin Time (APTT) test among subjects of African descent in Sokoto, North Western Nigeria. Samples of 99 subjects made up of 49 male and 50 female subjects with mean age 38.3 ± 22.3 years. Coagulation tests were performed immediately specified times after phlebotomy up to 24 hours (0, 1, 2, 3, 4 and 24 hours at room temperature of 40 degrees C. Our data demonstrate that prothrombin time and APTT results are stable for up to 2 hours, remaining constant regardless of storage conditions. Post hoc tests using Bonferroni correction revealed that there were increases in PT time from 0 hour to 4 hours (17.82 ± 0.61 seconds vs 18.30 ± 0.59 seconds, respectively), from 0 hour to 24 hours (17.82 ± 0.61 seconds vs 18.48 ± 0.59 seconds, respectively), from 2 hours to 4 hours (17.89 ± 0.58 seconds vs 18.30 ± 0.59 seconds), from 2 hours to 24 hours (17.89 ± 0.58 seconds vs 18.48 ± 0.58 seconds), which were all statistically significant (p = 0.002 and p < 0.000, p < 0.000, p < 0.000, respectively). However, the increase in PT time from 0 hour to 2 hours (17.82 ± 0.61 seconds vs 17.89 ± 0.59 seconds, respectively) and from 4 hours to 24 hours (18.30 ± 0.59 vs 18.48 ± 0.59 seconds, respectively) were not statistically significant (p = 1, p = 0.428). A repeated measure ANOVA determined that mean PTTK time differed statistically significantly between time points F (3, 291) = 119.22, p < 0.001. Post hoc tests using Bonferroni correction revealed that there were increase in PTTK time from 0 hour to 2 hours (37.86 ± 1.04 seconds vs 39.94 ± 1.07 seconds, respectively), from 0 hour to 4 hours (37.86 ± 1.04 seconds vs 42.34 ± 1.11 seconds, respectively), from 0 hours to 24 hours (37.86 ± 1.04 seconds vs 44.93 ± 1.20 seconds), from 2 hours to 4 hours (39.94 ± 1.07 seconds vs 42.34 ± 1.11 seconds), from 2 hours to 24 hours (39.94 ± 1.07 seconds vs 44.93 ± 1.20 seconds) and from 4 hours to 24 hours (42.43 ± 1.11 vs 44.93 ± 1.20 seconds), which were all statistically significant at p < 0.001). Therefore, we conclude that there are no statistically significant differences in the PT and APTT between 0 and 2 hours. A longer timing (after 2 hours) from phlebotomy collection of blood from respondents elicited a statistically significant increase in the PT and APTT result. There were no statistically significant differences in the PT and APTT result determined 4 hours and 24 hours after phlebotomy. Longer timing from collection of blood from respondents elicited a statistically significant increment/increase in the clotting time using PTTK. Our data demonstrate that PT and APTT results are stable for 2 hours remaining constant regardless of storage conditions.

Honey Bee Venom (Apis mellifera) Contains Anticoagulation Factors and Increases the Blood-clotting Time.

Objectives:Bee venom (BV) is a complex mixture of proteins and contains proteins such as phospholipase and melittin, which have an effect on blood clotting and blood clots. The mechanism of action of honey bee venom (HBV, Apis mellifera) on human plasma proteins and its anti-thrombotic effect were studied. The purpose of this study was to investigate the anti-coagulation effect of BV and its effects on blood coagulation and purification.Methods:Crude venom obtained from Apis mellifera was selected. The anti-coagulation factor of the crude venom from this species was purified by using gel filtration chromatography (sephadex G-50), and the molecular weights of the anti-coagulants in this venom estimated by using sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Blood samples were obtained from 10 rabbits, and the prothrombin time (PT) and the partial thromboplastin time (PTT) tests were conducted. The approximate lethal dose (LD) values of BV were determined.Results:Crude BV increased the blood clotting time. For BV concentrations from 1 to 4 mg/mL, clotting was not observed even at more than 300 seconds, standard deviations (SDs) = ± 0.71; however, clotting was observed in the control group 13.8 s, SDs = ± 0.52. Thus, BV can be considered as containing anti-coagulation factors. Crude BV is composed 4 protein bands with molecular weights of 3, 15, 20 and 41 kilodalton (kDa), respectively. The LD50 of the crude BV was found to be 177.8 μg/mouse.Conclusion:BV contains anti-coagulation factors. The fraction extracted from the Iranian bees contains proteins that are similar to anti-coagulation proteins, such as phospholipase A2 (PLA2) and melittin, and that can increase the blood clotting times in vitro.

Titanium oxide films with vacuum thermal treatment for enhanced hemocompatibility

Blood compatibility is a key property of biomaterials contacting blood. Titanium oxide film has been deposited on Si wafers (110) by unbalanced reactive magnetron sputtering and successive vacuum annealing. The structure, chemical composition, hydrophilicity and surface energy of the films have been investigated by X-ray diffraction, X-ray photoelectron spectroscopy and contact angle measurement respectively. The results revealed that high vacuum annealing increased the surface energy, hydrophilicity and the rate of of Lewis base in the film. In vitro evaluation of hemocompatibility, including platelet adhesion, fibrinogen denaturation, activated partial thromboplastin time and thrombin time test, proved that the hydrophilic titanium oxide films have excellent blood compatibility.

Investigation of adverse effects of interactions between herbal drugs and natural blood clotting mechanism.

Throughout the world, herbal medicines are consumed by most of the patients without considering their adverse effects. Many herbal medicines/plant extracts have been reported to interact with the natural blood clotting system. In continuation to this effort, thirty medicinal plant extracts were allowed to interact with citrated human blood and the clotting time was measured after re-calcification in vitro using Lee and White method. The aq. leaf ext. of Syzygium cumini and Camellia sinensis significantly prolonged the clotting time. In response to the prothrombin time and activated partial thromboplastin time tests, the ext. of C. sinensis showed normal APTT and marginally prolonged the PT to 16.7 s (control-15.2 s) while S. cumini showed normal PT but significantly prolonged the APTT to 66.9 s (control-20.7 s). This suggests that, C. sinensis acts on the extrinsic pathway while S. cumini on the intrinsic pathway. There are some common herbal formulations that are frequently used by the patients which contain above plant materials, like, Syzygium cumin in anti-diabetic formulations, while the ext. of C. sinensis is consumed frequently as beverage in many part of the world. Hence, patients having known bleeding tendency or haemophilia disease should take into account the interaction potential of these plants with the natural blood clotting system while taking herbal formulations containing above plants; specially, the patients suffering from intrinsic pathway factor deficiency should keep a limit on the consumption of S. cumini while extrinsic pathway factor deficiency patients should limit C. sinensis. Also, the medical practitioners should consider the patient's food consumption history before doing any major surgical procedures.

Evaluation of the main coagulation tests in the presence of hemolysis in healthy subjects and patients on oral anticoagulant therapy.

Our study was designed to evaluate, on healthy subjects and patients on oral anticoagulant therapy vitamin K antagonist (OAT-vka), the possible interference caused by hemolysis on the main coagulation tests. To obtain hemolyzed samples, two methods were used: heat shock and mechanical system. The coagulation tests on hemolyzed samples were performed employing optical automated analyser BCSxp (Siemens Healthcare(®)). Moreover, the prothrombin time (PT) and activated partial thromboplastin time (aPTT) tests were also carried out manually using an electromechanical device (KC4 - Amelung). The PT test, on healthy subjects, in case of moderate hemolysis can be performed without significant interference on automatic instrument. On manual instrument, the PT test can be performed even in case of marked hemolysis. For patients on OAT-vka, the PT test in case of marked hemolysis can be performed both on automatic and manual instrument. For the aPTT test, it can be carried out manually, because also in case of marked hemolysis a statistically significant difference was not observed. For the fibrinogen test, a dramatic concentration decrease was already clear for weak hemolysis. A decreased function on antithrombin test was statistically significant for mild-moderate hemolysis. The D-dimer test showed increased values for mild hemolysis. The rejection of hemolyzed sample and/or the request of a second sample are not always the proper attitudes to take for performing clotting tests. The rational management of the hemolyzed samples decreases the employment of both nursing and technical staff significantly, the turnaround time and, consequently, does not lead to additional costs for each patient involved.

Prothrombin Time and Activated Partial Thromboplastin Time Testing: A Comparative Effectiveness Study in a Million-Patient Sample.

BackgroundA substantial fraction of all American healthcare expenditures are potentially wasted, and practices that are not evidence-based could contribute to such waste. We sought to characterize whether Prothrombin Time (PT) and activated Partial Thromboplastin Time (aPTT) tests of preoperative patients are used in a way unsupported by evidence and potentially wasteful.Methods and FindingsWe evaluated prospectively-collected patient data from 19 major teaching hospitals and 8 hospital-affiliated surgical centers in 7 states (Delaware, Florida, Maryland, Massachusetts, New Jersey, New York, Pennsylvania) and the District of Columbia. A total of 1,053,472 consecutive patients represented every patient admitted for elective surgery from 2009 to 2012 at all 27 settings. A subset of 682,049 patients (64.7%) had one or both tests done and history and physical (H&P) records available for analysis. Unnecessary tests for bleeding risk were defined as: PT tests done on patients with no history of abnormal bleeding, warfarin therapy, vitamin K-dependent clotting factor deficiency, or liver disease; or aPTT tests done on patients with no history of heparin treatment, hemophilia, lupus anticoagulant antibodies, or von Willebrand disease. We assessed the proportion of patients who received PT or aPTT tests who lacked evidence-based reasons for testing.ConclusionsThis study sought to bring the availability of big data together with applied comparative effectiveness research. Among preoperative patients, 26.2% received PT tests, and 94.3% of tests were unnecessary, given the absence of findings on H&P. Similarly, 23.3% of preoperative patients received aPTT tests, of which 99.9% were unnecessary. Among patients with no H&P findings suggestive of bleeding risk, 6.6% of PT tests and 7.1% of aPTT tests were either a false positive or a true positive (i.e. indicative of a previously-undiagnosed potential bleeding risk). Both PT and aPTT, designed as diagnostic tests, are apparently used as screening tests. Use of unnecessary screening tests raises concerns for the costs of such testing and the consequences of false positive results.

Anti-inflammatory, anticoagulant and antioxidant effects of aqueous extracts from Moroccan thyme varieties

ObjectiveTo evaluate the anti-inflammatory, anticoagulant and antioxidant effects of aqueous extracts of thyme varieties from Moroccan. MethodsThe aqueous extracts of tree medicinal plants [Thymus atlanticus (T. atlanticus), Thymus satureioides and Thymus zygis (T. zygis)] were screened for their antioxidant activity using 1,1-diphenyl-2-picrylhydrazyl radical-scavenging, ferric reducing antioxidant power assay, radical scavenging activity method, the inhibition of 2,2'-azobis (2-amidinopropane) dihydrochloride that induces oxidative erythrocyte hemolysis and thiobarbituric acid reactive substances assay. The anti-inflammatory activity of aqueous extracts was evaluated in vivo using croton oil-induced ear edema and carrageenan-induced paw edema in mice and rats, respectively. This extracts were evaluated in vitro for their anticoagulant activity at the different concentrations by partial thromboplastin time and prothrombin time activated. ResultsAll thyme varieties were found to possess considerable antioxidant activity and potent anti-inflammatory activity in the croton oil-induced edema. Administration of aqueous extracts of two varieties (50 mg/kg) (T. zygis and T. atlanticus) reduced significantly the carrageenan-induced paw edema similar to non-steroidal anti-inflammatory drug (indomethacin, 10 mg/kg). In partial thromboplastin time and prothrombin time tests, T. atlanticus and T. zygis extracts showed the strongest anticoagulant activity. In contrast, Thymus satureioides did not show the anticoagulant activity in these tests. ConclusionsAll aqueous extracts possess considerable antioxidant activity and are rich in total polyphenol and flavonoid but they act differently in the process of inflammatory and coagulation studied. This study shows great variability of biological activities in thyme varieties.

Biochemical characterization of prothrombin complex concentrates in China

Despite increasing use of prothrombin complex concentrates (PCCs), there is little knowledge about the biochemical characterization of Chinese PCCs. Six Chinese PCCs were investigated and compared with PCCs (Octaplex®) from Europe. The levels of coagulation factors and inhibitors were detected. The presence of activated coagulation factors was assessed. Furthermore, their thrombin inhibitory capacities, specific activity and purity were assayed. All above parameters of biochemical properties were statistically analyzed.Chinese PCCs contained FⅡ, Ⅶ, Ⅸ and Ⅹ, protein C, S and Z, heparin and extremely low level antithrombin, as well as Octaplex®. The measured FⅨ activities were similar to those declared, however the measured potency of FⅡ, Ⅶ and Ⅹ greatly exceeded the labeled. Though all preparations were negative for activated coagulation factors in non-activated partial thromboplastin time test, the activated coagulation factor Ⅶ (FⅦa) remained in all PCCs and its content differed greatly. Overall, FⅦa content of Chinese PCCs was higher than that of Octaplex®. Further, Chinese PCCs were inferior to Octaplex® in the thrombin inhibitory capacities, specific activity and purity. In summary, compared with Octaplex®, Chinese PCCs' errors about the labeled activity of coagulation factors and probably high risks of thrombosis should be considered.

Validation Studies on Coagulation Laboratory Assays Useful for Assessing the Plasma Levels of Apixaban

Apixaban(BMS/Pfizer), is clinically developed for DVT prophylaxis in patients undergoing hip and knee surgery, stroke prevention in patients with atrial fibrillation, and the prevention of acute ischemic events in patients with acute coronary syndrome. As claimed for other new oral anticoagulants (dabigatran, rivaroxaban), there appears to be no need to routinely monitor apixaban. However, there are clinical settings that would benefit from monitoring (e.g., patients who do not fit the enrollment criteria of those patients evaluated through the clinical trials). There are also clinical settings where assessing drug levels would be a critical component of patient care (e.g., patients admitted to the Emergency Department; stroke patients; patients on drug with failed anticoagulant protection or with bleeding; patients on drug who require interventional procedures or surgery, etc.). At this time a laboratory assay for apixaban has not yet been defined. Data from several laboratories demonstrates that the prothrombin time (PT) and activated partial thromboplastin time (aPTT) tests do not provide adequate sensitivity to apixaban at typical clinically obtained or higher concentrations. This was consistently found with multiple different manufacturers’ reagents for both the PT and aPTT. This collaborative study was designed to provide information on potential laboratory tests that maybe suitable for the clinical assessment of plasma levels of apixaban.Material & Methods: Five coagulation laboratories were included in this collaborative study. Each laboratory received an apixaban standard at 1.0 ug/ml, normal human pooled plasma and unknown samples labeled A, B, C, D and E. Each day for 5 days, a standard 6 point curve in the range of 0-1.0 ug/ml and samples A-E were run in duplicate using the Technoclone anti-Xa assay (Technoclone, Vienna, Austria), Coamatic anti-Xa assay (Chromogenix; Instrumentation Laboratory; Bedford, MA) and the Prothrombin time using Innovin Siemens, Deerfield, IL). All date was sent to the Hemostasis & Thrombosis Research Laboratory, Loyola University Chicago for analysis.Results: The measured apixaban levels as measured by various methods are shown in the following Table. TableSampleTechnochromCoamaticInnovin PT1000 ng/ml1051.50±48.2 ng/ml968.26±38.6 ng/ml362.06±138.5 ng/ml500 ng/ml525.1±32.5 ng/ml540.85±40.9 ng/ml944.09±245.4 ng/ml250 ng/ml239.3±28.8 ng/ml267.2±29.7 ng/ml468.5±21.60 ng/ml125 ng/ml140.4±24.6 ng/ml191.16±27.01 ng/ml678.55±147.8 ng/ml30 ng/ml25.5±8.9 ng/ml38.7±14.2 ng/ml155.19±93.3 ng/mlThe Technochrom anti-Xa and Coamatic anti-Xa assays were the most accurate in determining the concentration of apixaban in the plasma. There was a large standard deviation which was due to the variation in instrumentation between the laboratories. Each laboratory adapted the three assays to their current coagulation analyzers. The PT overestimated the apixaban concentration at <500 ng/ml and under estimated at higher concentration.Conclusions: These studies suggest that the anti-Xa chromogenic methods from Technoclone and Chromogenix may be used to determine the concentration of apixaban in plasma. Each method needs to be validated on the specific coagulation instrument to be used to assess the apixaban concentration. DisclosuresNo relevant conflicts of interest to declare.

Direct oral anticoagulants: new drugs and new concepts.

Direct oral anticoagulants (DOACs) are approved for multiple thromboembolic disorders and provide advantages over existing agents. As with all anticoagulants, management protocols for the eventuality of bleeding are important. Randomized phase III studies generally show that DOACs have a similar risk of clinically relevant bleeding compared with standard anticoagulants, with reductions in major bleeding in some cases. This may be particularly important in patients with atrial fibrillation, for whom the rate of intracranial hemorrhage was approximately halved with DOACs compared with warfarin. Conversely, the risk of gastrointestinal bleeding may be increased. Specific patient characteristics, such as renal impairment, comedications, and particular aspects of each drug, including the proportion eliminated by the kidneys, must be taken into account when assessing the risk of bleeding. Although routine coagulation monitoring of DOACs is not required, it may be useful under some circumstances. Of the traditional clotting assays, a sensitive and calibrated prothrombin time may be useful for detecting the presence or absence of clinically relevant factor Xa inhibitor concentrations (rivaroxaban or apixaban), but specific anti-factor Xa assays can measure drug levels quantitatively. For dabigatran, the results of an activated partial thromboplastin time test may exclude a clinically relevant pharmacodynamic effect, but a calibrated dilute thrombin time assay can be used for quantification of drug levels. In the event of mild or moderate bleeding, normal hemostatic support measures are recommended. For life-threatening bleeding, use of nonspecific prohemostatic agents may be considered, although clinical evidence is scarce. Specific antidotes are in development.