D. Kristol

The effect of ultrasound on the alkaline hydrolysis of nitrophenyl esters

Abstract Ultrasound was found to increase the rate of hydrolysis of a series of esters by up to 15%. No effect of molecular structure upon this enhancement was observed.

Effect of endotoxin and silver ion on the clotting time of blood

Endotoxin (Lipopolysaccharide, LPS) is a part of the outer membrane of the cell wall of gram-negative bacteria. LPS induces activation of the coagulation cascade in humans. This procoagulant effect on citrated whole blood produces a shortened clotting time. Silver ion (Ag/sup +/) denatures the anticoagulant proteins and affects the intrinsic pathway of blood coagulation by producing a shortened clotting time. This in vitro study was performed by mixing human citrated whole blood (CWB) with LPS, silver nitrate (AgNO/sub 3/), and the combination of the two. The clotting time, rate of clot formation, and the maximum strength of the clot were determined. The results show that the combination of LPS and Ag/sup +/ significantly reduces the clotting time (p<.002).

In vitro investigation of streptokinase activity using Sonoclot coagulation analysis

Streptokinase (SK) therapy has become a common clinical intervention in the treatment of acute coronary syndromes. Recent clinical trials indicate that adjuvant treatment with low molecular weight heparin (LMWH) or bivalirudin (BV) may attenuate fibrinolytic-induced procoagulant activity. We investigated the effects of SK with LMWH and BV on the clotting of whole blood using a Sonoclot/sup /spl reg// Coagulation Analyzer. Human citrated whole blood was spiked with SK and LMWH at final concentrations of control, 5 U/mL SK, 0.2 U/mL LMWH, and 5 U/mL SK:0.2 U/mL LMWH. The samples (n=10) were incubated at 37/spl deg/C for 10 minutes and recalcified with CaCl/sub 2/ to initiate clotting. Tracings were obtained using the Sonoclot Signature Viewer/spl trade/ software, and the clotting time (CT) and clot viscosity were noted. The experiment was repeated (n=8) using 5 /spl mu/g/mL BV in place of LMWH. The Sonoclot tracings provide a whole-picture analysis of the effects of SK and adjuvant anticoagulants on haemostatic and fibrinolytic mechanisms. Implementation of coagulation monitoring protocols for SK therapy may aid in clinical diagnosis.

A functional clotting assay to monitor low molecular weight heparin dosage

Low molecular weight heparin (LMWH), an anticoagulant drug, has seen increased use in a variety of clinical situations. Mercuric ion (Hg/sup 2+/) has been shown to increase procoagulant activity via Tissue Factor (TF) stimulated production of Factor Xa, a pathway that has been linked to many thrombotic disorders. The effects of Hg/sup 2+/ on heparinized blood may provide valuable insight into the effectiveness of LMWH to counteract TF-induced hypercoagulability. We performed in vitro studies by spiking human citrated whole blood (CWB) with HgCl/sub 2/ and LMWH at final concentrations of 0.01 % HgCl/sub 2/, 0.5 U/mL LMWH, and a combination of these agents. The samples (n=19) were incubated at 37/spl deg/ for 10 minutes and recalcified with 0.1 M CaCl/sub 2/ to initiate clotting. The clotting or recalcification time (RT) was measured with a Sonoclot Coagulation Analyzer. The results indicate that HgC12 was able to significantly (p<0.0001) reduce the anticoagulant ability of LMWH. The data warrants a reassessment of current LMWH monitoring and therapeutic protocols.

A computer assisted receptor mapping approach to the design of anti-AIDS agents directed at HIV reverse transcriptase

The authors take a pharmacophoric approach to the identification of a substrate/inhibitor binding site on HIV-1 RT. A pharmacophore is defined as the spatial arrangement of a set of atoms or groups in a ligand molecule while bound to a given receptor. The pharmacophores usefulness is in the assumption that a single one exists for a series of compounds binding at the same receptor site. The superposition of a series of substrate and/or inhibitors along a pharmacophoric pattern provides an enzyme excluded volume, which represents a minimum volume requirement for substrate/inhibitors at the receptor binding site. The structure of HIV-1 RT complexed with dsDNA template-primer has been recently described at 7 /spl Aring/ resolution and is currently being extended to 3 /spl Aring/. The combination of crystallographic, pharmacophoric, and existing biochemical and genetic data regarding the substrate binding residues will help to generate an accurate structure of the substrate binding domain of HIV-1 RT and aid in the evaluation of new and specific RT inhibitors.<<ETX>>

Thrombus formation and blood contacting surface in pneumatic diaphragm blood pump

For investigating the thrombus formation in a pneumatic diaphragm blood pump and the critical factors responsible for thrombus formation on smooth polyurethane blood contacting surfaces, four in vivo implanted blood pumps (implanted for 6 to 25 days) and four unused pumps were studied by scanning electron microscopy (SEM). The results demonstrate that one of the important reasons for thrombus formation in blood pumps is the morphology of the surface inside the blood chamber. Some surface defects were observed on the blood contacting surface. The defects could have been caused by material properties, the fabrication process, or excessive bending stress. The diaphragm and housing junction (DHJ) is a critical area where the thrombus formation of ten occurs. Marked reduction in thrombus formation for an improved pump is attributed to material pretreatment, stringent quality control, and reasonable redesign of the DHJ structure.<<ETX>>

Effect of tissue factor on plasma and packed RBC clotting parameters

Tissue factor (TF) is a transmembrane glycoprotein and the main triggering element for blood coagulation. Expression of TF on the cell surfaces and its appearance as a soluble molecule are characteristic features of acute and chronic inflammation in conditions such as arteriosclerosis leading to stroke and myocardial infection. A study was performed on human citrated whole blood, by separating plasma from the red blood cells. Ten microlitres of 25% tissue factor (TF) was added to both the plasma and packed red blood cells (RBCs), and the clotting time, rate of clot formation and the maximum strength of the clot determined. TF bearing samples clotted more rapidly and had greater clot strength when compared to that of control aliquots.

The effect of copper ion on blood coagulation

The effect of copper ions on the blood coagulation time was determined. In the experiment that was performed in vitro, there were three different aliquots for each blood sample. The three aliquots were composed of the control, 0.05 mM CuSO/sub 4/, and 0.1 mM CuSO/sub 4/. Thirty two microliters of 0.1 M CaCl/sub 2/ was then added to each aliquot to initiate blood coagulation. When the copper ion concentration was increased, there was a significant increase in blood coagulation time.

In vitro investigation of the procoagulant of silver ion

Silver ion, a potent antimicrobial and procoagulant, is hypothesized to achieve its haemostatic effect by binding anticoagulant proteins. We performed in vitro studies to gain more insight into its mechanism of action. 10 /spl mu/L of 5% AgNO/sub 3/ was added to 1 mL human plasma (n = 4), forming a precipitate. The precipitate was recovered through centrifugation and iteratively washed with saline. The supernatant(s) and precipitate were mixed with Coomassie blue stain and protein content was measured via spectrophotometric absorbance. An analysis of variance indicated significant (p<0.0001) differences between all groups. Multiple comparison tests showed a significant (p<0.001) increase in absorbance from the final wash and dye blank to the precipitate, indicating the presence of proteins in the precipitate. Silver ion may be precipitating anticoagulant proteins as its mechanism of action.

The effect of copper ions on sedimentation rates of erythrocytes

Whether copper sulfate affects the sedimentation rate of erythrocytes in blood was investigated. We performed the experiment by placing by 1ml of blood in plastic vials. After that, we added 10/spl mu/l of 1% CuSO/sub 4/ solution to an aliquot. Then, we put 10/spl mu/l of 5% of CuSO/sub 4/ to 1ml of the same blood into different vial. Following this procedure, we incubated the blood with the various concentrations of CuSO/sub 4/ to 37 degrees Celcius for ten minutes. Then, we determined the sedimentation rates of the red blood cells (RBCs) for the control, the blood that contained .001% of CuSO/sub 4/, and the blood that contained .005% of CuSO/sub 4/. We repeated this experiment with fourteen (n=14) different samples of blood. The results indicate that there was a statistical significance (P<.05) in the sedimentation rate of the blood that contained the .005% of CuSO/sub 4/ and the control were different In addition, there was a statistical significance (P<.05) that there was a difference in sedimentation rate between the .001% CuSO/sub 4/ and .005% CuSO/sub 4/. The results also show that there is no difference in sedimentation between .001% and the control.