Combination of ultrasound-assisted ethyl chloroformate derivatization and switchable solvent liquid-phase microextraction for the sensitive determination of l-methionine in human plasma by GC-MS.

A green and fast analytical method for the determination of l-methionine in human plasma is presented in this study. Preconcentration of the analyte was carried out by switchable solvent liquid phase microextraction after ethyl chloroformate derivatization reaction. Instrumental detection of the analyte was performed by means of gas chromatography-mass spectrometry. N,N-Dimethyl benzylamine was used in the synthesis of switchable solvent. Protonated N,N-dimethyl benzylamine volume, volume/concentration of sodium hydroxide, and vortex period were meticulously fixed to their optimum values. Besides, ethyl chloroformate, pyridine, and ethanol volumes were optimized in order to get high derivatization yield. After the optimization studies, limit of detection and quantitation values were attained as 3.30 and 11.0 ng/g, respectively, by the developed switchable solvent liquid phase microextraction gas chromatography-mass spectrometry method that corresponding to 76.7-folds enhancement in detection power of the gas chromatography-mass spectrometry system. Applicability and accuracy of the switchable solvent liquid phase microextraction-gas chromatography-mass spectrometry method were also checked by spiking experiments. Percent recovery results were ranged from 97.8 to 100.5% showing that human plasma samples could be analyzed for its l-methionine level by the proposed method.

Accurate, sensitive determination of omega-6 and omega-3 polyunsaturated fatty acids in human plasma, urine samples.

Quantitative determination of omega-6 and omega-3 polyunsaturated fatty acids in human plasma and urine with high accuracy and precision provides significant information to monitor the underlying etiology of several diseases. In this regard, liquid chromatography-mass spectrometry is a good choice owing to its great selectivity and sensitivity. Additionally, the hybrid quadrupole-time of flight-mass spectrometer systems provides easy identification of target compounds with superior mass measurements. In this study, an analytical method has been developed for simple, accurate and simultaneous determination of linoleic acid, arachidonic acid, docosahexaenoic acid and eicosapentaenoic acid in a short chromatographic analysis period. The developed method is suitable for the quantitative detection of these four compounds with detection limits ranging between 1.1-3.0 ng ml-1 and its applicability was assessed in human urine and plasma samples. As a result, acceptable accuracy (between 83 and 111%) and good precision (<6%) were obtained for target compounds using matrix matching calibration strategy.

Accurate and sensitive determination of sildenafil, tadalafil, vardenafil, and avanafil in illicit erectile dysfunction medications and human urine by LC with quadrupole-TOF-MS/MS and their behaviors in simulated gastric conditions.

The widespread use of phosphodiesterase-5 inhibitors has attracted broad attention of counterfeiters to develop illicit erectile products with inaccurate amounts, unknown toxicity, and purity of active ingredients. Correspondingly, intake of these products endangers consumer health and needs to be screened for precautionary actions to reduce this risk. Therefore, in this study, a sensitive and rapid analytical method has been developed for simultaneous determination of selected phosphodiesterase-5 inhibitors present in illicit erectile medications and human urine. Quantification of the analytes was performed by liquid chromatography coupled with quadrupole-time-of-flight tandem mass spectrometry system. The chromatographic separation was successfully achieved with a run period of 8 min. Low detection limits were obtained in the range of 1.63-9.81 ng/g with relative standard deviations below 7.72% obtained using the replicate measurements of lowest concentration in calibration plots. The analytical performance of the proposed method proved good linearity, low detection limits, good accuracy and precision with high percent recoveries for human urine samples. Developed method was successfully applied to real samples including four different brands of illicit erectile medications. The results obtained revealed the presence of high levels of sildenafil in analyzed samples. The behaviors of selected phosphodiesterase-5 inhibitors were also studied in simulated gastric conditions.

Magnetic cobalt particle-assisted solid phase extraction of tellurium prior to its determination by slotted quartz tube-flame atomic absorption spectrophotometry.

The emergence of magnetic materials has opened up doors to numerous applications including their use as sorbents for preconcentration of trace elements. Magnetic materials exhibit many unique advantages in sample preparation such as easy separation from the sample, high preconcentration factor, and short operation period. In the present study, magnetic cobalt material was synthesized, characterized, and used as an effective sorbent in a solid phase extraction process. Experimental variables of the extraction process including pH and volume of buffer solution, eluent concentration and volume, mixing type and period, and sorbent amount were optimized to achieve maximum extraction efficiency. Instrumental variables of flame atomic absorption spectrophotometry and the type of slotted quartz tube were also investigated. Under the optimum conditions, the combined method provided a wide linear range between 50 and 200 ng/mL with detection and quantification limits of 15.4 ng/mL and 51.3 ng/mL, respectively. Relative standard deviations of the proposed method were less than 5.0% and a high enrichment factor of 86.7 was obtained. The proposed method was successfully applied to soil samples for the determination of trace tellurium.

A novel analytical approach for the determination of parathion methyl in water: quadrupole isotope dilution mass spectrometry-dispersive liquid-liquid microextraction using multivariate optimization.

Dispersive liquid-liquid microextraction was coupled with quadruple isotope dilution mass spectrometry for the sensitive and accurate determination of parathion methyl in water. The two methods were complementary to each other, with DLLME preconcentrating the analyte for trace determination, and ID4MS maintaining the integrity of the method's accuracy and precision. An experimental design was used to optimize the extraction process. The results from the design were evaluated with the analysis of variance to determine the statistical significance of the main factors of extraction and interaction effects of these factors. A three-point calibration blend and sample blend were prepared gravimetrically by spiking with isotopically labelled parathion methyl. All four blends were left to equilibrate for three hours, after which they were preconcentrated under the optimum extraction conditions. The percent recovery recorded by this method was 99.9%, and the percent relative standard deviation was 0.32%. These results validated the accuracy and precision of the combined method.

Simultaneous determination of niacin and pyridoxine at trace levels by using diode array high-performance liquid chromatography and liquid chromatography with quadrupole time-of-flight tandem mass spectrometry.

A highly sensitive and simple diode-array high-performance liquid chromatography and liquid chromatography with quadrupole time-of-flight tandem mass spectrometry method was developed for the simultaneous determination of niacin and pyridoxine in pharmaceutical drugs, tap water, and wastewater samples. To determine the in vivo behavior of niacin and pyridoxine, analytes were subjected to simulated gastric conditions. The calibration plots of the diode-array high-performance liquid chromatography and liquid chromatography with quadrupole time-of-flight tandem mass spectrometry method showed good linearity over a wide concentration range with close to 1.0 correlation coefficients for both analytes. The limit of detection/limit of quantitation values for liquid chromatography quadrupole time-of-flight tandem mass spectrometry analysis were 1.98/6.59 and 1.3/4.4 µg/L for niacin and pyridoxine, respectively, while limit of detection/limit of quantitation values for niacin and pyridoxine in high-performance liquid chromatography analysis were 3.7/12.3 and 5.7/18.9 µg/L, respectively. Recovery studies were also performed to show the applicability of the developed methods, and percentage recovery values were found to be 90-105% in tap water and 94-97% in wastewater for both analytes. The method was also successfully applied for the qualitative and quantitative determination of niacin and pyridoxine in drug samples.

Nonuraemic nonfatal idiopathic calciphylaxis in a kitten.

BACKGROUND: Calciphylaxis is a rare cutaneous disorder, characterized by vascular calcification and progressive skin necrosis, not yet described in cats. It is scarcely reported in animals, mostly due to iatrogenic or uraemic disturbances of the calcium-phosphate balance. In human patients, it is most commonly seen with end-stage renal disease, but several nonuraemic disorders, including inherited dysfunctions of tissue calcification inhibitors, have also been described. HYPOTHESIS/OBJECTIVES: To describe a case of nonuraemic calciphylaxis in a cat. ANIMALS: A 10-week-old male domestic short hair kitten was presented with hyperacute skin lesions. Initial dermatological signs were characterized by sharp demarcated erosions and ulcerations on the face, including the nasal planum and lips. Cutaneous lesions rapidly progressed into thick crusts with ulcerations, involving parts of the face and pinna as well as abdominal skin. METHODS: Complete blood count, serum chemistry profile, urinalysis, parathyroid hormone measurement and histopathological examination of skin biopsies. RESULTS: Histopathology from newly developed abdominal skin lesions revealed severe epidermal necrosis and calcification, multifocal pannicular calcification and calcified subcutaneous vessels, supporting a diagnosis of calciphylaxis. Treatment consisted of systemic and topical antimicrobials, analgesics, pentoxifylline, Lantharenol(®) , sodium thiosulfate and vitamin K. After initiation of therapy, no further progression was noticed; all medications could be discontinued eventually, and no relapse was seen in the following 2 years. CONCLUSIONS AND CLINICAL IMPORTANCE: Calciphylaxis should be considered as a differential diagnosis for ulcerative cutaneous disorders in young cats. More information on this disease is needed to elucidate the pathomechanism.

Pulmonary hypertension in systemic lupus erythematosus: pulmonary thromboembolism is the leading cause.

BACKGROUND: Pulmonary hypertension (PH) is a life-threatening complication of systemic lupus erythematosus (SLE). Pulmonary hypertension in SLE has a variety of causes. Diagnosing early and defining the cause of PH accurately can provide better clinical outcome in SLE. We investigated the causes and characteristics of PH in patients with SLE. METHODS: One hundred twenty-one patients with SLE who had a visit in a 6-month period were assessed retrospectively. Patients who ever had a systolic pulmonary arterial pressure of 40 mm Hg or greater by Doppler echocardiography were considered to have PH. RESULTS: Among 122 patients, 65 had echocardiography for some reason, and 10 (8.2%) were diagnosed as having PH by echocardiographic examination. This number reduced to 9 (7.4%) when we excluded the patient with normal pulmonary artery pressure at right heart catheterization. Causes of PH were as follows: thromboembolic events in 4 patients (44.4%) (2 of them had chronic thromboembolic PH), left-sided heart disease in 2 patients (22.2%), pulmonary arterial hypertension in 1 patient (11.1%), high cardiac output state in 1 patient (11.1%), and transient elevation of systolic pulmonary artery pressure in 1 patient (11.1%) who had a history of venous thromboembolism. Venous thromboembolic disease was significantly higher in patients with SLE with PH in comparison to patients with SLE without PH (7 patients [6.3%] vs 5 patients [50.0%]; P = 0.001). All patients improved clinically during their short-term follow-up. CONCLUSIONS: Patients with SLE are at increased risk for PH. This study highlights the complexity of the differential diagnosis of PH in patients with SLE once again and emphasizes the importance of pulmonary thromboembolism as a cause of PH. One should investigate patients with SLE with unexplained symptoms and/or signs related to PH for possible treatable causes.

Development of an analytical method for the determination of pesticides in tropical fruits by LC-QTOF-MS/MS after QuEChERS extraction sample cleanup and DLLME preconcentration.

In this study, QuEChERS extraction was combined with dispersive liquid-liquid microextraction (DLLME) to extract pesticides from tropical fruits for determination by a highly accurate and sensitive liquid chromatography-quadrupole time-of-flight mass spectrometry (LC-QTOF-MS/MS) system. The QuEChERS method served as a matrix clean-up tool and the DLLME method preconcentrated the analytes for their determination at trace levels. All parameter variables of the DLLME method were optimized to improve the extraction output and lower the limits of detection and quantification (LOD and LOQ) for all the analytes. Under the optimum experimental conditions, the LOD and LOQ values were found in the range of 0.004-0.013 and 0.27-0.61µg l-1, respectively. The detection limits achieved by direct LC-QTOF-MS/MS analysis were increased by about 10-260 folds using the optimized DLLME method. To assess the accuracy and applicability of the developed method, spike recovery experiments on tropical fruits were carried out. The matrix matching calibration method was used to enhance the quantification accuracy of the analytes in kiwi, pineapple, and mango matrices, with percent recoveries ranging between 89 and 117%.

Magnetic Nanoparticles Based Solid Phase Extraction Methods for the Determination of Trace Elements.

The past three decades has seen significant advancements in instrumental detection techniques for both organic and inorganic analytes, leading to more accurate and precise measurements. The relevance of sample preparation prior to instrumental reading cannot be underestimated, and this area has also seen major improvements, with solid phase extraction (SPE) and its miniaturized version (solid phase microextraction, SPME) serving as an efficient clean-up and preconcentration method for analytes. Magnetic nanoparticles (MNPs) belong to the new generation of sorbents, and their characteristic small size and easy manipulation under magnetic field make them ideal materials for extraction of several analytes. This review presents the basic principles of SPE and SPME, and lays more emphasis on different types of MNPs that have been used as efficient separation and preconcentration tools. In addition, application of MNPs for the determination of trace elements in environmental, biological and food samples are well documented in this review.

Accurate determination of amino acids by quadruple isotope dilution-reverse phase liquid Chromatography-Tandem mass spectrometry after derivatization with 2-Naphthoyl chloride.

The determination of amino acids in biological samples is central to the diagnosis of inherited metabolic disorders and also gives significant information about the metabolisms in the cells and living body. The development of analytical method for reliable quantification of amino acids in biological samples is still challenging because of the polar nature of amino acids and complex nature of biological samples causing a high degree of interferences during analysis. In the present study, a pre-column derivatization method using 2-naphtoyl chloride combined with liquid chromatography-tandem mass spectrometry method was developed for the determination of 17 amino acids in human serum and urine matrices. Low detection limits were obtained in the range of 0.015 - 0.266 µmol kg-1 and acceptable recovery results were obtained in human serum and urine samples. Isotopically labelled (15N labelled) amino acids were spiked to standards and samples before derivatization to compensate for the analytical errors in the whole procedure. The combination of quadrupole isotope dilution strategy with the derivatization based reversed phase chromatography allowed to improve method accuracy and precision.

Serum Levels of Selected Elements in Patients with Beta Thalassemia Major.

ß-thalassemia major is a significant health problem in the world that obliges patient to repeated blood transfusions. Frequent transfusions cause toxic iron overload and the association between serum iron levels and ß-thalassemia major have been extensively studied in literature. Nonetheless, data on trace elements is still limited. The aim of this work was to study the relationship between some trace element levels in serum and ß-thalassemia major. The quantifications of Al, K, Sn, Sb, Ni, Cr, Cd, Ba, Co, As and Se elements were carried out by Inductively Coupled Plasma - Mass Spectrometry system. Mann Whitney U test is performed in order to test the statistical difference between patient and control groups in terms of their element concentrations. Significant differences were observed for the concentrations of Al, K, Sn and Sb elements and for the correlation between concentrations of K-Sb elements. The study indicates higher levels of Al and Sb, and lower levels of K and Sn elements of patients when compared to control group. These findings reveal the altered profile of serum trace element concentrations and so, further studies are required to evaluate the potential of trace elements as biomarkers and/or to administrate their levels in blood to reduce the related complications.

Influence of blood collection technique on platelet function and coagulation variables in dogs.

OBJECTIVE: To investigate the influence of diameter of the catheter and blood collection technique on platelet function and variables reflecting secondary hemostasis, physiologic anticoagulation, and fibrinolysis in dogs. ANIMALS: 6 healthy Beagles. PROCEDURES: Blood samples were collected with 20- and 18-gauge venous catheters immediately after catheters were inserted in a peripheral vein, through a 14-gauge central venous catheter that had been placed via the Seldinger technique in a jugular vein < 30 minutes before sample collection, and through a 13-gauge central venous catheter placed via a catheter-through-the-needle technique < 30 minutes before sample collection (techniques 1 to 4, respectively). Platelet function was assessed in hirudin-anticoagulated whole blood via an impedance-based aggregometer with collagen (0.8, 0.4, 0.2, 0.1, and 0.05 µg/mL) as an inductor. Kaolin-activated thromboelastography variables were determined in citrated whole blood. Prothrombin time, activated partial thromboplastin time, fibrinogen and fibrin D-dimer concentrations, and activity of factor VIII, antithrombin, protein C, and protein S were assessed automatically in citrated plasma. RESULTS: At 0.05 µg of collagen/mL, the highest median rate of aggregation was observed for collection techniques 2 and 3 with 4.3 (range, 2.5 to 6.5) and 3.7 (range, 2.8 to 8.3) aggregation units/min; however, these values were not significantly different from values for the other collection techniques. Generally, sample collection technique did not have a significant impact on results of coagulation variables investigated. CONCLUSIONS AND CLINICAL RELEVANCE: Various blood collection techniques can be used to obtain samples for coagulation testing.

Determination of seventeen free amino acids in human urine and plasma samples using quadruple isotope dilution mass spectrometry combined with hydrophilic interaction liquid chromatography - Tandem mass spectrometry.

Taking into account the growing demand for new analytical procedures that are appropriate for analysis of complex biological samples with increased sensitivity, accuracy and precision, a novel analytical method was described for the determination of underivatized amino acids in human plasma and urine samples. The presented analytical procedure involved the direct analysis of urine samples and the analysis of plasma samples followed by a simple protein precipitation protocol. Samples were analyzed using a simple and fast chromatographic method developed for the determination of 17 different amino acids by liquid chromatography - tandem mass spectrometry. The limit of detection and quantification values for amino acids were ranged between 0.03-2.26 µmol kg-1 and 0.09-7.54 µmol kg-1. Matrix effects of plasma and urine on the quantification of analytes were determined by spiking experiments. The accuracy of method was evaluated by matrix matching and quadruple isotope dilution strategies. Excellent accuracy and precision were obtained with the use isotope labeled amino acids demonstrating the high reliability and reproducibility of the proposed method. The percent recovery values were found to be between 98.70 - 101.68% with%RSD below than 1.62% for human plasma and 99.14 - 101.78% with%RSD below than 2.44% for urine samples.

Sodium, Magnesium, Calcium, Manganese, Iron, Copper, and Zinc in Serums of Beta Thalassemia Major Patients.

Thalassemia major is the most severe form of thalassemia and occurs with the impaired synthesis of ß-globin which causes the accumulation of unpaired alpha globin chain. Patients with beta thalassemia major can only survive with periodically safe blood transfusions leading to the accumulation of iron in the bloods of patients, and this causes several endocrinopathies. Although iron overload in thalassemic patients has been extensively studied, there is little information about the levels of other trace elements. The aim of this study was to investigate the differences of serum concentrations of sodium, magnesium, calcium, manganese, iron, copper, and zinc for patients with major ß-thalassemia. Concentration of elements was determined by inductively coupled plasma-mass spectrometry system. The statistical difference between the elemental concentrations of the patient and control groups was found by the Mann-Whitney U test. In addition, the relationship between concentrations of the measured elements for each group was determined by the Spearman correlation test. The results revealed that iron, zinc, magnesium, and manganese serum levels of thalassemic patients were significantly higher than the control group while calcium concentration was statistically lower than the control group. There was no significant difference observed for copper and sodium levels of patients when compared to the healthy control group.

An accurate and sensitive analytical method for the simultaneous determination of glycine, methionine and homocysteine in biological matrices by matrix matching strategy and LC-quadrupole-time-of-flight-MS/MS.

Nowadays, it is well known that early diagnosis directly affects the success of treatment. Biomarkers play a crucial role in early diagnosis of diseases or explanation of pathological condition. The investigation of new biomarkers depends on the reliable quantification of analytes in biological matrices. Regarding to the critical roles of amino acids in metabolism, functions and nutrition of human body, the careful monitoring of their levels in biological samples is required to evaluate their potential in biomarker studies for clinical research. In this study, a reliable and accurate analytical strategy was developed for the simultaneous determination of glycine, methionine and homocysteine using LC-quadrupole-time of flight-tandem MS system. The method detection limit was found to be 0.73 µg/mL, 0.017 µg/mL and 0.019 µg/mL for glycine, methionine and homocysteine, respectively. The calibration curves were obtained with great linearity (R2 ≥ 0.9993) and low relative standard deviation values showed the repeatability of proposed method. The method applicability was determined using human plasma and urine samples, and high percent recoveries demonstrated the accuracy of method developed. Each measurement was taken less than 4.0 min indicating a promising strategy for the fast and reliable quantification of target amino acids in clinical laboratories.

A novel determination method for diuron in seaweed samples: Combination of quadruple isotope dilution strategy with liquid chromatography - quadrupole time of flight - tandem mass spectrometry for superior accuracy and precision.

This paper proposed an accurate and sensitive analytical method based on the combination of Liquid Chromatography-Quadrupole Time of Flight-Tandem Mass Spectrometry (LC-QTOF-MS/MS) system with Quadruple Isotope Dilution-Mass Spectrometry (ID4MS) strategy for the determination of diuron at trace levels. In order to achieve accurate and reliable determination of the analyte, ID4MS strategy that uses stable isotopically labelled analogues was employed. Diuron-d6 was synthesized in the laboratory using a novel strategy and employed in ID4MS for the preparation of three calibration blends and seaweed sample blend. The analytical performance of the LC-QTOF-MS/MS method was evaluated and the respective detection and quantification limits obtained were 14.6 µg kg-1 and 46.5 µg kg-1. Good linearity was obtained with a correlation coefficient of 0.9995 over the dynamic range from 50 to 1000 µg kg-1. The validity of the method was successfully tested by carrying out spiking experiments in seaweed samples. The percent recovery value showed superior enhancement for ID4MS strategy (99.97 ± 0.41%) confirming the applicability of the method to complex matrices with great accuracy and precision.

Trace determination of cobalt in biological fluids based on preconcentration with a new competitive ligand using dispersive liquid-liquid microextraction combined with slotted quartz tube-flame atomic absorption spectrophotometry.

A new competitive ligand has been synthesized for the preconcentration to obtain lower detection limits by using dispersive liquid-liquid microextraction combined with slotted quartz tube-flame atomic absorption spectrophotometry (DLLME-SQT-FAAS). The proposed method is simple, eco-friendly and has high sensitivity. The preconcentration procedure was optimized on the basis of various parameters affecting the complex formation and extraction efficiency such as pH and volume of buffer solution, volume of ligand solution, mixing period, volume and type of extraction solvent, volume and type of dispersive solvent, and salt effect. Instrumental parameters were also optimized to get higher sensitivity. Under the optimum conditions, the calibration graph was linear in the range of 10-250 ng mL-1and the resulted limits of detection and quantification (LOD and LOQ) for combined method were 4.7 and 15.7 ng mL-1, respectively. The detection power was improved 48-fold using DLLME-SQT-FAAS method compared to conventional FAAS. The precision of the method was found to be high with a relative standard deviation of 2.5%. The accuracy of method was evaluated by recovery experiments using matrix matching study on spiked urine and blood samples. The recoveries for urine and blood samples ranged from 99.8 to 108.9% and 102.5 to 110.0%, respectively.

The Relationship Between the Surface Morphology and Chemical Composition of Gunshot Residue Particles.

In this study, chemical composition and morphology of gunshot residue (GSR) of 9 × 19 mm Parabellum-type MKE (Turkey)-brand ammunition were analyzed by scanning electron microscope and energy dispersive X-ray spectrometer. GSR samples were collected by "swab" technique from the shooter's right hand immediately after shooting. According to general principles of thermodynamics, it is likely that the structures will have a more regular (homogeneous) spherical form to minimize their surface area due to very high temperatures and pressures that occur during explosion. Studied samples were collected under the same conditions with the same original ammunition, from the same firearm and a single shooter. This is because many other variables may affect size, structure, and composition in addition to the concentrations of elements of the structure. Results indicated that the chemical compositions are effective in the formation of GSR morphological structures.

Effect of submaximal aerobic exercise on platelet function, platelet activation, and secondary and tertiary hemostasis in dogs.

OBJECTIVE: To investigate whether submaximal aerobic exercise in dogs is followed by activation of all phases of coagulation as has been reported for humans. ANIMALS: 9 healthy Beagles. PROCEDURES: 30 minutes before dogs were exercised, a 16-gauge central venous catheter was placed in a jugular vein of each dog by use of the catheter-through-the-needle technique. Samples were collected before exercise, after running on a treadmill (6 km/h for 13 minutes), and at 60 minutes. Platelet activation was evaluated with platelet morphology indices (mean platelet component, mean platelet volume, and number of large platelets) provided by a laser-based hematology system. Platelet function was assessed in hirudin-anticoagulated whole blood with an impedance-based aggregometer with collagen as the agonist (final concentrations, 0, 1.6, 3.2, 5, and 10 µg/mL). Prothrombin time, activated partial thromboplastin time, and concentrations of fibrinogen, factor VIII, antithrombin, protein C, protein S, and fibrin D-dimer were determined automatically. Kaolin-activated thromboelastography variables R (reaction time), K (clot formation time), angle α, maximal amplitude, and G (clot stability) were measured in recalcified citrated whole blood. RESULTS: Exercise resulted in a significant decrease in mean platelet volume and the number of large platelets but did not change the mean platelet component, which reflected platelet activation as well as platelet function. Secondary and tertiary coagulation did not change significantly, nor did thromboelastography variables. CONCLUSIONS AND CLINICAL RELEVANCE: Aerobic exercise resulted in a decrease in the number of large and thus most likely activated platelets but otherwise had no major impact on coagulation in dogs.