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Saudi Pharmaceutical Journal : SPJ logoLink to Saudi Pharmaceutical Journal : SPJ
. 2013 Oct;21(4):405–410. doi: 10.1016/j.jsps.2012.12.001

A new and simple HPLC method for determination of etamsylate in human plasma and its application to pharmacokinetic study in healthy adult male volunteers

Sally A Helmy a,⁎, Heba M El Bedaiwy b
PMCID: PMC3824948  PMID: 24227961

Abstract

A new and simple HPLC assay method was developed and validated for the determination of etamsylate in human plasma. After protein precipitation with 6% perchloric acid, satisfactory separation was achieved on a HyPURITY C18 column (250 mm × 4.6 mm, 5 μm) using a mobile phase comprising 20 mM sodium dihydrogen phosphate-2 hydrate (pH was adjusted to 3.5 by phosphoric acid) and acetonitrile at a ratio of 95:5 v/v. The elution was isocratic at ambient temperature with a flow rate of 0.75 ml/min. Allopurinol was used as internal standard. The calibration curve was linear over the range from 0.25 to 20 μg/ml (r2 = 0.999). The limit of quantification for etamsylate in plasma was 0.25 μg/ml. The within day coefficient of variance (%CV) ranged from 3.9% to 10.2%, whereas the between-day %CV ranged from 3.1% to 8.7%. The assay method has been successfully used to estimate the pharmacokinetics of etamsylate after oral administration of a 500 mg tablet under fasting conditions to 24 healthy Egyptian human male volunteers. Various pharmacokinetic parameters including AUC0–t, AUC0–∞, Cmax, Tmax, t1/2, MRT, Cl/F, and Vd/F were determined from plasma concentration–time profile of etamsylate.

Keywords: Etamsylate, HPLC, Validation, Plasma, Pharmacokinetics

1. Introduction

Etamsylate is an orally administered hemostatic drug. It is used in the treatment of capillary hemorrhage, hematemesis, hemoptysis, malena, hematuria, epistaxis, menorrhagia and post partum hemorrhage (Sweetmann, 2002). It works by increasing capillary endothelial resistance and promoting platelet adhesion. It reduces capillary bleeding in the presence of normal number of platelets. In addition, etamsylate is effective in the prevention and treatment of pre-, per-, or postsurgical capillary hemorrhages in all delicate operations and in those affecting highly vascularized tissues such as E.N.T., gynecology, obstetrics, urology, ophthalmology, plastic and reconstructive surgery (Symes, 1975; Harrison and Campbell, 1976). Etamsylate is chemically 2,5-dihydroxy benzene sulfonic acid with N-ethylethanamine (Fig. 1). The molecular formula of etamsylate is C10H17NO5S and the molecular weight is 263.33 g/mol. It is completely soluble in water, methanol, and ethanol but partially soluble in methylene chloride (Merck Index, 1989).

Figure 1.

Figure 1

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Chemical structure of etamsylate.

Several analytical methods were developed for the assay of etamsylate in pharmaceutical formulations. These methods include; spectrophotometry (Xu et al., 1994; El-Shabrawy et al., 2004; El-Enany et al., 2007; Goyal and Singhvi, 2008; Zhang et al., 2010), HPTLC (Jaiswal et al., 2005a,b), HPLC (Ma and Liu, 1984; Xu et al., 1994; El-Shabrawy et al., 2004; Kaul et al., 2005; Jaiswal et al., 2005a,b; El-Enany et al., 2007; Nagaraju et al., 2008; Goyal and Singhvi, 2008; Zhang et al., 2010; Chandrshekhara et al., 2010; Vamshikrishna and Sathish Kumar, 2011), electrochemical methods (Wang et al., 2001,2005; Wang and Xu, 2007; Zhang and Wang, 2005), chemiluminescence (Zhang et al., 1998; Yang et al., 2002; Li et al., 2002; Du et al., 2002) and capillary electrophoresis (Li and Ju, 2006). To the best of our knowledge, no HPLC method has been yet reported for the determination of etamsylate in human plasma. Also, no pharmacokinetic studies have been published before. Owing to its great therapeutical importance and widespread use, studying the pharmacokinetic parameters of the drug is a prerequisite for the current study. Therefore, a new and simple validated HPLC assay method for the determination of etamsylate in human plasma was developed.

2. Materials and methods

2.1. Chemicals

Etamsylate and allopurinol were purchased from Sigma–Aldrich Co., St. Louis, MO, USA; Dicynone® tablets (500 mg Etamsylate) were purchased from OM Pharma, Geneva, Switzerland; All chemicals and reagents used were of LC grade and were purchased from Merck Chemicals, Germany; Milli-Q grade (Millipore, Bedford, MA, USA) water was used in all cases. Serum was prepared from normal human blood, procured freshly from healthy volunteers through a local blood bank and pooled for spiking and using as control blank samples.

2.2. Instruments

The analysis of etamsylate was performed in HPLC system equipped with a variable-wavelength ultraviolet (UV) detector and an automatic injector (LC-10 VP, Shimadzu Scientific Instrument, Kyoto, Japan). Data analysis program Class-VP (Shimadzu Scientific Instruments, Kyoto, Japan) was used for data acquisition and processing. Other instruments used in this study include Bath sonicator (UltraSonik 57X, Elmsford, USA); Balance (Sartorius, Germany); Magnetic stirrer (Stuart, Stone, Staffordshine, UK); Centrifuge (Hettich, Germany); pH meter (520 laboratory instrument, USA); Vortex tube mixer (Heidolph, Germany); Brand auto micro-pipettes (Labnet, USA); and micro liter syringes from Hamilton (Bonaduz, Switzerland).

2.3. HPLC system and operating conditions

Separation was accomplished with a HyPURITY C18 column (250 mm × 4.6 mm, 5 μm; Thermo Fisher Scientific, Waltham, MA). The mobile phase comprised 20 mM sodium dihydrogen phosphate-2 hydrate (pH was adjusted to 3.5 by phosphoric acid) and acetonitrile at a ratio of 95:5 v/v; the elution was isocratic at ambient temperature with a flow rate of 0.75 ml/min. The UV detector was set at 309 nm for etamsylate and 254 for the internal standard (IS; allopurinol). The detector was programmed at 254 nm for 6.8 min and at 309 nm for the subsequent 1.2 min to measure etamsylate and IS, respectively, and the peak areas were calculated using the data analysis program Class-VP (Shimadzu Scientific Instruments, Kyoto, Japan).

2.4. Standard solutions

For the construction of calibration curves, drug stock solution (500 μg/ml) was prepared by dissolving etamsylate powder in methanol. The stock solution was further diluted in methanol to prepare seven working solutions of etamsylate with concentrations of (200, 100, 50, 20, 10, 5 and 25 μg/ml) for spiking human plasma samples. Standard solutions were stored at +4 °C. The internal standard stock solution was prepared by dissolving allopurinol powder in methanol to give concentration of 30 μg/ml which is used as the IS for etamsylate analysis in human plasma.

2.5. Plasma sample preparation

The calibration standards of etamsylate were prepared by transferring 25 μl from each working solution of etamsylate and IS to a set of test tubes. The solvent was evaporated, and 0.25 ml of blank plasma was added to each tube to form a set of calibration standards with concentrations of 0.25, 0.5, 1, 2, 5, 10 and 20 μg/ml. The sample was vortexed, and then 0.25 ml of 6% trichloroacetic acid was added, shaken for one min, and centrifuged at 3000 rpm for 7 minutes. The supernatant was separated and 25 μl was injected into the HPLC.

2.6. Assay validation

The method was validated with respect to linearity, selectivity, precision, accuracy, specificity, and stability.

2.6.1. Linearity

The linearity of the calibration curve in human plasma was confirmed by plotting the peak-area ratios of (drug/IS) versus the corresponding etamsylate concentration with least-squares linear regression analysis.

2.6.2. Selectivity

The retention time of the drug in the standard curve of spiked plasma blanks and the study samples were compared to define any endogenous materials and/or degradation peaks appeared at the same retention time of etamsylate and IS in the HPLC chromatograms.

2.6.3. Precision and accuracy

The precision of the assay was measured by the within day and between-day percent coefficient of variation (%CV) over the concentration range of calibration curve of etamsylate in plasma during the course of validation. Within day precision of the assay was determined by the analysis of three replicates of the calibration curve in the same day. The between-day precision was evaluated by the analysis of six different calibration curves in six different days during the study period. The %CV of the measured concentrations was used to determine the precision of the assay. The accuracy of the assay was defined as the absolute mean value of the ratio of the back calculated mean values of the unknown samples and their nominal values, expressed as a percentage. The measured concentrations of the within day and between-day analysis were compared to the nominal concentrations to determine the accuracy of the assay.

2.6.4. Specificity

Three quality control samples (QC) at concentrations of 0.5, 2.0 and 10.0 μg/ml, (LLOQ, MQC and HQC) were prepared as unknown samples. The concentration in each QC sample was determined from the calibration curve and the calculated concentration was compared with the nominal concentration. Intra-day precision and accuracy were evaluated by analyzing each QC sample six times on the same day, while inter-day precision and accuracy were evaluated by analyzing each QC sample in 6 consecutive days.

2.6.5. Stability

The stability of the drug at room temperature was determined by preparing three different plasma samples for each drug concentration then the samples were injected immediately into the HPLC. The samples were kept at room temperature and were injected again after 24 h. The concentrations measured at time zero and after 24 h were compared to determine any changes in the concentrations with time. The 24 h period was selected because it is the maximum time between the preparation of samples and injection into the HPLC. The stability of the drug in frozen plasma was investigated by the analysis of the study samples obtained from three volunteers twice. The first analysis was performed at the beginning of the study and the second analysis was performed at the end of the study. The samples were stored at −80 °C between the analyses.

2.7. Clinical pharmacokinetic study

2.7.1. Study design

This study was monitored in accordance with the International Conference of Harmonisation (ICH) guidance on general considerations for clinical trials.

2.7.2. Study population

Twenty-four male Egyptian volunteers with a mean ± SD age of 22.70 ± 4.090 years (range, 18–32 years) and mean ± SD weight of 69.37 ± 7.037 kg (range, 58–82 kg) were chosen to participate in the present study. All subjects completed an acceptable medical history, medication history, physical examination, an electrocardiogram, blood pressure, screens for HIV-1 and -2 antibodies and hepatitis B surface antigen, and a urine drug screen prior to study initiation. Selected routine clinical laboratory measurements were performed during screening. Exclusion criteria were: (1) treatment with any known enzyme-inducing/inhibiting agents within 30 days prior to the start of the study and throughout the study; (2) susceptibility to allergic reactions to drugs; (3) any prior surgery of the gastrointestinal tract that may interfere with drug absorption. Upon completion of study, the physical examination and clinical laboratory measurements were repeated. The subjects were instructed to abstain from taking any medication for 1 week prior to and during the study period. All enrolled volunteers were healthy, and none of the participants showed any signs of adverse drug reactions during or after completion of the study. Informed consent was obtained from the subjects after explaining the nature and purpose of the study. The study protocol was approved by the ethics committee of Tanta University Hospital, Egypt.

2.7.3. Dosing schedule

In the morning of study, after an overnight fasting (10 h), volunteers received a single dose of one Dicynone® tablet (500 mg etamesylate) followed by 200 ml of water. The volunteers were continuously monitored by a qualified staff throughout the confinement period of study. Food and drinks were withheld for at least 2 h after dosing. Water intake was allowed after 2 h of dose; water, lunch and dinner were given to all volunteers according to a time schedule. All subjects were asked to abstain from the consumption of fruit juices during the study period.

2.7.4. Collection of blood samples

Venous blood (5 ml) was taken from the forearm by vein puncture prior to (0 h) and at 0.25, 0.5, 0.75, 1, 1.5, 2, 2.5, 3, 4, 5, 6, 8, 10, 12, 24 and 48 h after each drug intake. The blood samples were drawn into 10 ml heparinized tubes and plasma was directly separated by centrifugation at 3000 rpm for 10 min, transferred into polypropylene plastic tubes, and stored at −80 °C until assay.

2.7.5. Monitoring of adverse effects

Throughout the study, the subjects were questioned and examined for the presence of adverse drug reactions. All the participated volunteers did not show any sign of adverse drug reactions during or after completing the study.

2.7.6. Determination of pharmacokinetic parameters

The developed HPLC method was used to investigate the plasma concentration–time profile of etamsylate after administration of 500 mg tablet. Study samples were treated as the calibration standards. A non-compartmental model was used to determine the pharmacokinetic parameters of etamsylate. The elimination rate constant (Ke) was estimated by least squares regression of plasma concentration–time data points in the terminal log-linear region of the curves. Half-life (t1/2) was calculated as 0.693 divided by Ke. The AUC from zero to the last quantifiable plasma concentration (AUC0–t) was calculated using the linear trapezoidal rule. The AUC from zero to infinity (AUC0–∞) was calculated as AUC0-∞ (AUC0–t + C/Ke), where C is the last measured concentration (Gilbaldi and Perrier, 1982; Hedaya, 2007). Cmax and tmax were obtained directly from the individual plasma concentration–time curve. The apparent total clearance (Cl/F) was calculated as oral clearance (CL/F) = Dose/AUC0–∞ while the apparent volume of distribution (Vd/F) was estimated as Oral Clearance/K relative to the bioavailability (F) of etamsylate. Area under the first moment curve (AUMC) was calculated by trapezoidal integration and extrapolation to infinity. Mean residence time (MRT) was calculated as the ratio (AUMC)/(AUC0–∞) (Gilbaldi and Perrier, 1982; Hedaya, 2007).

3. Results and discussions

3.1. Conditions of chromatography

Method development was focused on the development and optimization of suitable sample preparation and chromatographic separation. Several tests were performed for optimizing the components of mobile phase in order to achieve good chromatographic peak shape and resolution. Good separation of etamsylate and IS with short run times were obtained using a mobile phase system of 20 mM sodium dihydrogen phosphate-2 hydrate (pH was adjusted to 3.5 by phosphoric acid) and acetonitrile at a ratio of 95:5 v/v; at 0.75 ml/min flow rate. The UV detector was programed at 309 nm for 6.8 min and at 254 nm for the subsequent 1.2 min to measure etamsylate and IS, respectively at retention times that were quite short for IS and etamsylate at 6.3 ± 0.09 and 7.2 ± 0.1 minutes, respectively. For plasma samples, deproteinization with 6% trichloroacetic was adopted, which was simple and fast resulting in sharp and symmetrical peaks.

3.2. Validation of HPLC method

3.2.1. Linearity

Calibration curves were linear with a correlation coefficient R2 > 0.999 throughout the course of the assay (20–0.25 μg/ml) for etamsylate as shown in Fig. 2.

Figure 2.

Figure 2

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Calibration curve of etamsylate in spiked plasma (mean ± S.D.; n = 8).

3.2.2. Selectivity

Typical chromatograms obtained from blank human plasma and the plasma spiked with etamsylate and IS are shown in Fig. 3, which indicates that no peaks for any endogenous compound appeared at the same retention time for etamsylate and IS.

Figure 3.

Figure 3

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Representative chromatograms for blank plasma sample and blank plasma spiked with etamsylate to produce a drug concentration of 10 μg/mL and the internal standard (IS).

3.2.3. Precision and accuracy

Precision and accuracy results demonstrated good precision and accuracy over the concentration range selected. Intra- and inter-day precision and accuracy for etamsylate from plasma data are listed in Table 1. The within day and between-day %CV were always within ±10% in the entire range of the calibration curve. The within day %CV ranged from 3.9% to 10.2%, whereas, the between-day %CV ranged from 3.1% to 8.7%. The intra-day accuracy ranged between 99.0% and 109.4%, whereas the inter-day accuracy ranged between 98.4% and 108.2%.

Table 1.

The day-to-day (intra-day) and between-days (inter day) precision and accuracy of the assay.

Nominal concentration (μg/ml) Day-to-day (intra-day)a
 Between-days (inter-day)b
Concentration found (μg/ml) (mean ± S.D) Precision (%) Accuracy (%) Concentration found (μg/ml) (mean ± S.D) Precision (%) Accuracy (%)
20 20.9 ± 1.21 5.77 104.8 20.09 ± 1.27 6.32 100.4
10 10.2 ± 0.40 3.91 102.3 10.14 ± 0.32 3.15 101.4
5 5.10 ± 0.32 6.38 102.1 5.185 ± 0.40 7.81 103.7
2 1.98 ± 0.20 10.2 99.00 1.97 ± 0.08 4.41 98.5
1 1.08 ± 0.07 6.67 109.4 0.984 ± 0.04 4.33 98.4
0.5 0.50 ± 0.03 6.09 101.8 0.541 ± 0.04 8.53 108.2
0.25 0.27 ± 0.01 4.44 108.0 0.251 ± 0.02 8.76 100.4
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a

n = 3 for each concentration.

b

n = 6 for each concentration.

3.2.4. Specificity

Intra- and inter-day precision and accuracy for the three quality control samples in human plasma were evaluated and depicted in Table 2.

Table 2.

Quality control samples.

Nominal concentration (μg/ml) Concentration found (μg/ml; mean ± S.D.) Precision (%) Accuracy (%)
10 9.84 ± 0.99 10.0 98.4
2.0 1.89 ± 0.13 6.87 94.5
0.5 0.53 ± 0.05 9.39 106.4
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3.2.5. Stability

Short-term stability showed that etamsylate is stable in plasma for at least 24 hrs at room temperature. The difference in the drug concentration in all samples obtained from volunteers in the two analyses performed at the beginning and at the end of the study was always less than 10% in each sample in frozen plasma stored at −80 °C during the study period, indicating the stability of etamsylate in frozen plasma.

The described analytical method was proven to be sensitive and accurate for the determination of etamsylate in plasma where, all validation parameters were within the acceptable limits which indicate the suitability of the developed analytical method for pharmacokinetic study.

3.3. Pharmacokinetic results

The developed method was used for the pharmacokinetic study of etamsylate in 24 healthy Egyptian volunteers. After oral administration of 500 mg etamsylate, the concentration versus time profile of etamsylate was constructed. The mean plasma concentration–time curve of etamsylate is shown in Fig. 4. The results showed that the method was reliable and adequate to provide pharmacokinetic concentration–time profile for a dose of etamsylate of 500 mg tablet. Etamsylate was readily absorbed from the gastrointestinal tract and it was measurable at the first sampling time (0.25 h) in almost all volunteers. The major pharmacokinetic parameters of etamsylate were listed in Table 3.

Figure 4.

Figure 4

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Mean plasma concentrations of etamsylate in 24 healthy volunteers after a single oral dose administration of one Dicynone® tablet (500 mg etamsylate) (mean ± standard error).

Table 3.

Pharmacokinetic parameters of etamsylate after a single oral dose administration of one Dicynone® (500 mg etamsylate) to 24 healthy Egyptian male volunteers.

Parameters Etamsylate
Mean %CV
Cmax (μg/ml) 8.05 30.3
tmax (h) 5.02 27.7
AUC0 → t (μg h/ml) 78.7 32.6
AUC0 → ∞ (μg h/ml) 84.8 31.0
t1/2 (h) 9.4 34.9
MRT (h) 15.7 14.8
Cl/F (ml/min) 6470.5 25.1
Vd/F (ml) 86629.7 23.4
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Cmax, peak plasma concentration; tmax, time to reach peak plasma concentration; AUC0–∞, area under the concentration–time curve from zero to infinity; AUC0–t, area under the concentration–time curve from zero to the last measurable plasma concentration; AUCt–∞, area under the concentration–time curve from the last measurable plasma concentration to infinity; t1/2, elimination half-life; MRT, mean residence time; Cl/F, clearance; Vd/F, volume of distribution.

4. Conclusions

A new and simple HPLC method for the determination of etamsylate in human plasma was developed and validated over the concentration range from 0.25–20 μg/ml. The method has been successfully applied to the pharmacokinetic study of etamsylate in Egyptian healthy male volunteers and satisfactory results were obtained, which demonstrates that the method is reproducible, sensitive and reliable. Owing to great therapeutical importance of etamsylate and widespread use; this work is considered as a primary step in preparing controlled release preparations in the future.

Footnotes

Peer review under responsibility of King Saud University.

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References

  1. Chandrshekhara S., Prakash G., Shamrez A., Basavaraj K., Mahibub M., Yusrida D. A new stability-indicating RP-HPLC method to determine etamsylate in injection formulation. Int. J. Pharm. Biol. Arch. 2010;1(2):140–143. [Google Scholar]
  2. Du J., Li Y., Tang Y., Lu J. Flow-injection chemiluminescence determination of etamsylate based on permanganate oxidation. Anal. Lett. 2002;35(3):463–472. [Google Scholar]
  3. El-Enany N., Belal F., Rizk M. Kinetic spectrophotometric determination of etamsylate in dosage forms. J. AOAC Int. 2007;90(3):679–685. [PubMed] [Google Scholar]
  4. El-Shabrawy Y., El-Enany N., Salem K. Sensitive kinetic spectrophotometric determination of captopril and etamsylate in pharmaceutical preparations and biological fluids. Farmaco. 2004;59(10):803–808. doi: 10.1016/j.farmac.2004.05.005. [DOI] [PubMed] [Google Scholar]
  5. Gilbaldi, M., Perrier, D., 1982. Pharmacokinetics. In: Marcel Dekker (Ed.), 2nd ed., Informa Healthcare Inc., New York, USA.
  6. Goyal A., Singhvi I. Spectrophotometric estimation of etamsylate and mefenamic acid from a binary mixture by dual wavelength and simultaneous equation methods. Indian J. Pharm. Sci. 2008;70(1):108–111. doi: 10.4103/0250-474X.40345. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Harrison R., Campbell S. A double-blind trial of etamsylate in the treatment of primary and intra-uterine induced menorrhagia. Lancet. 1976;2:283–285. doi: 10.1016/s0140-6736(76)90733-9. [DOI] [PubMed] [Google Scholar]
  8. Hedaya M.A. In: Taylor and Francis, editor. Taylor and Francis group; 6000 Broken Sound Parkway NW, Suit 300, Boca Raton, FL 33487-2742, England: 2007. (Basic pharmacokinetics). [Google Scholar]
  9. Jaiswal Y., Talele G., Surana S. Quantitative analysis of etamsylate and mefenamic acid in tablets by use of planar chromatography. J. Planar. Chromatogr. - Mod. TLC. 2005;18(106):460–464. [Google Scholar]
  10. Jaiswal Y.S., Talele G., Surana S. A simple and sensitive HPTLC method for quantitative analysis of etamsylate in tablets. J. Planar. Chromatogr. Mod. TLC. 2005;105(18):380–383. [Google Scholar]
  11. Kaul N., Agrawal H., Kakad A., Dhaneshwar S., Patil B. Stress degradation studies on etamsylate using stability-indicating chromatographic methods. Anal. Chim. Acta. 2005;536(1–2):49–70. [Google Scholar]
  12. Li J., Ju H. Simultaneous determination of etamsylate, tramadol and lidocaine in human urine by capillary electrophoresis with electrochemiluminescence detection. Electrophoresis. 2006;27(17):3467–3474. doi: 10.1002/elps.200600135. [DOI] [PubMed] [Google Scholar]
  13. Li Y., Du J., Lu J. Chemiluminescence reaction of luminol-[potassium ferricyanide-potassium ferrocyanide]-etamsylate system. Fenxi Huaxue. 2002;30(6):742–744. [Google Scholar]
  14. Ma J., Liu Y. Quantitative analysis of etamsylate by h.p.l.c. Yaowu Fenxi Zazhi. 1984;4(4):209–211. [Google Scholar]
  15. Nagaraju P., Krishnachaithanya K., Srinivas V.D.N., Padma S.V.N. Reverse phase high performance liquid chromatographic method for the determination of etamsylate in bulk and tablet dosage forms. Asian J. Res. Chem. 2008;1(2):88–90. [Google Scholar]
  16. Sweetmann S.C. Martindale –The Complete Drug Reference. Pharmaceutical Press; London (UK): 2002. p. 732. [Google Scholar]
  17. Symes J. The effect of dicynene on blood loss during and after transurethral resection of the prostate. Br. J. Urol. 1975;47:203. doi: 10.1111/j.1464-410x.1975.tb03948.x. [DOI] [PubMed] [Google Scholar]
  18. The Merck Index . 11th ed. Merck; USA: 1989. An Encyclopedia of Chemicals, Drugs and Biologicals. (p. 3757) [Google Scholar]
  19. Vamshikrishna N., Sathish Kumar Shetty A. Development and Validation of RP-HPLC Method for the estimation of Etamsylate in Bulk drug and Pharmaceutical formulations. Int. J. ChemTech Res. 2011;3(2):928–932. [Google Scholar]
  20. Wang S., Xu Q. Electrochemical parameters of etamsylate at multi-walled carbon nanotube modified glassy carbon electrodes. Bioelectrochemistry. 2007;70(2):296–300. doi: 10.1016/j.bioelechem.2005.12.005. [DOI] [PubMed] [Google Scholar]
  21. Wang Z., Li Z., Zhou S. Voltammetric behavior of embedded super-thin carbon film electrode and its application. Fenxi Huaxue. 2005;33(4):523–526. [Google Scholar]
  22. Wang Z., Zhang D., Zhang Y., Zhou S. Voltammetric behaviour of Dicynone at a poly(4-aminopyridine) film-modified electrode and its determination by adsorptive stripping voltammetry. Fenxi Huaxue. 2001;29(1):83–86. [Google Scholar]
  23. Xu Z., Li X., Shi J., Yao X., Liu A., He Z. Determination of Dicynone [etamsylate] in gastric mucosa by ultra-violet spectrophotometry. Fenxi Huaxue. 1994;22(4):420. [Google Scholar]
  24. Yang F., Zhang C., Baeyens W., Zhang X. Determination of etamsylate in pharmaceutical preparations based on an auto-oxidation chemiluminescence reaction. J. Pharm. Biomed. Anal. 2002;30(3):473–478. doi: 10.1016/s0731-7085(02)00096-1. [DOI] [PubMed] [Google Scholar]
  25. Zhang C., Huang J., Feng M., Zhang Z. Flow injection chemiluminescence determination of etamsylate with electrogenerated hypochlorite. Anal. Lett. 1998;31(11):1917–1928. [Google Scholar]
  26. Zhang M., Zhang Y., Li Q. A novel visible spectrophotometric method for the determination of etamsylate in pharmaceutical preparations and biological samples. Spectrochim. Acta A. 2010;75(3):1013–1017. doi: 10.1016/j.saa.2009.12.039. [DOI] [PubMed] [Google Scholar]
  27. Zhang X., Wang S. Determination of etamsylate in the presence of catecholamines using 4-amino-2-mercaptopyrimidine self-assembled monolayer gold electrode. Sens. Actuators B. 2005;104(1):29–34. [Google Scholar]

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