The chromatographic determination of the chlorhexidine digluconate in the medical sponges (hemostatic sponges) Текст научной статьи по специальности «Фундаментальная медицина»

PHARMACEUTICAL SCIENCES

THE CHROMATOGRAPHIC DETERMINATION OF THE CHLORHEXIDINE DIGLUCONATE IN THE MEDICAL SPONGES (HEMOSTATIC SPONGES)

Pavliuk B.

I. Horbachevsky Ternopil National Medical University

Ternopil, Ukraine ChubkaM.

I. Horbachevsky Ternopil National Medical University

Ternopil, Ukraine Hroshovyi T.

I. Horbachevsky Ternopil National Medical University

Ternopil, Ukraine

ABSTRACT

A topical issue in modern pharmaceutical technology is the development of new drugs and expanding the range of available dosage forms in each pharmacological group. The development of the composition and technology of medical/hemostatic sponge involves the development of appropriate quality control techniques.

Therefore, the purpose of our work is to develop methods for the determination of digluconate chlorhexidine in the sponge composition by a modern, accurate and selective method of analysis.

High performance liquid chromatography identified and quantified the content of chlorhexidine digluconate in structured medical/hemostatic sponges, and developed appropriate methods of analysis. The obtained data can be recommended for the determination of the investigated active pharmaceutical ingredient in the composition of medicines.

Keywords: HPLC, chlorhexidine digluconate, identification, medical/hemostatic sponge

Actuality. Medical sponge - a tool that exhibits absorbent and antiseptic properties, as well as stimulates tissue regeneration. Absorption/hemostatic sponge are widely used in surgery, neurosurgery, dentistry, otolaryngology and gynecology as they help to stop blood loss or are used to close wound surfaces (burns, trophic ulcers). Sponge have bactericidal, antiseptic, antimicrobial, regenerating, tonic and absorbent properties and purposefully affect to the hearth of pathology.

The Food and Drug Administration (FDA) has published a retrospective review and identified absorb-able, collagen-based hemostatic agents that are potentially suitable and effective. Collagen-based hemostatic agents (medical sponge) can be particularly popular in the minimally invasive surgery, as a means of blood clotting in inaccessible places, or as a second line of defense when conventional methods are not sufficient to control bleeding. The FDA estimates that in 2012 year in more than 6.9 million procedures were used hemostatic agents. This figure summarizes 165 general related medical device reports from July 24, 2003 to December 31, 2018, inclusive.

Research and development of the topical hemo-static and absorptive agents is rapidly expanding since 2000, and most evidence confirms their effectiveness over the gauze.

Today, to obtain hemostatic/absorbent sponges can be used many polymeric materials. In particular, the technology of improvement biomedical sponges with natural biopolymers has attracted great attention from researchers. Commonly used absorbent hemo-static materials include collagen and collagen fibers, cellulose and hemicellulose, medical hemostatic gelatin, alginate, chitosan and others. Sponge as a versatile

form, which can be filled with the necessary active pharmaceutical ingredient (API), respectively they can be analgesic (with lidocaine, novocaine) or anti-inflammatory (with diclofenac, dimexide); hemostatic (with thrombin, calcium chloride, boric acid, calcium glu-conate) and with other API.

Today, we have developed a medical/hemostatic sponge based on a crushed cryoliophilized xenoderm substrate to stop bleeding and to close wound (burn) surfaces.

Chlorhexidine digluconate (C34H54Cl2N10O14), which belongs to cationic compounds and is one of the first internationally recognized therapeutic and prophylactic antiseptics for skin and wounds, was selected as an active pharmaceutical ingredient. Chlorhexidine di-gluconate is widely used in gynecology, surgery, urology, ophthalmology, otolaryngology and dental practice. One of the advantages of chlorhexidine diglu-conate, in addition to its pronounced antimicrobial action, is its ability to bind to various biological substrates and maintain its antibacterial activity and effective concentration in the composition of the drug. The mechanism of action of chlorhexidine digluconate based on the ability to alter the properties of cell membranes of microorganisms. The spectrum of antimicrobial activity spreads to bacteria, fungi of the genus Candida, enveloped viruses, long-term exposures to myco-bacteria and dermatophytes. Maximum efficiency is shown at pH values from 5.5 to 7.

Therefore, for the choice of therapeutic dosage of API (namely chlorhexidine digluconate) was guided by the data on the method of administration and dosage of drugs registered in Ukraine containing this API, the range of which is significant in the domestic pharma-

ceutical market. According to the instructions for medical use and review of literary sources, chlorhexidine digluconate recommended for use in doses of 0.05-2%. Accordingly, the dose of chlorhexidine digluconate per medical sponge was determined - 1%.

To date chlorhexidine digluconate does not belong to the State Pharmacopoeia of Ukraine, but there are relevant monographs in the European and American Pharmacopoeias, according to which it is proposed to use infrared absorption spectrophotometry for identification, as well as qualitative color and sedimentary reactions, for quantitative determination the method of liquid chromatography with spectrophotometric detection is proposed [8]. In particular, the authors used HPLC methods to develop a technique for quantifying chlorhexidine digluconate in vaginal suppositories. [9]. In the literary sources are given the examples of other methods (titrimetric, physicochemical) for quantitative determination of chlorhexidine digluconate in various dosage forms. In particular, the authors propose to use the method of capillary electrophoresis for the quantitative determination of chlorhexidine digluconate in 0.05 % solution, a spectrophotometric technique for the identification and quantitative determination of chlor-hexidine in dental drug films or titrimetric method to determine this API in solution, dental gel and developed appropriate methods of analysis [10-13].

The aim of our work was to confirm the possibility of using a modern, expressive and accurate method of High Performance Liquid Chromatography (HPLC) to determine the active pharmaceutical ingredient in medical sponges.

The purpose of the work was to develop a methodology for the identification and quantification of chlorhexidine digluconate in porous, structured sponges by HPLC.

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Fig. 1:

For the quantitative HPLC-determination of chlor-hexidine digluconate by the absolute calibration method, was used a calibration graph, constructed in the coordinates S, mm2 (peak area) - C, mg/ml (concentration of solutions of the test substance). For construct calibration graph using a standard solution of

Materials and methods:

Chromatographic analysis chlorhexidine digluconate performing on a liquid chromatography Agilent 1200 (Agilent technologies, USA) with diode-matrix detector.

The brief scheme of the experiment:

• Column: Zorbax SB-C18, size 2,1 mm*150 mm, 3,5 mkm;

• Mobile phase: A-ACN, B-0.5M phosphate buffer pH=3.0; 0,2 % triethylamine.

• Elution mode isocratic, the ratio (30/70, v/v).

• The rate of mobile phase: 0,5 ml/minutes;

• Column temperature: 40 °C;

• Wavelength of the diode-matrix detector: at a length of 245 nm, with the fixation of the spectrum in the range of 200-400 nm.

Reagents and chemicals:

Chlorhexidine digluconate solution 20 % in H2O was purchased from Sigma-Aldrich (USA), as well as certified auxiliaries of domestic and foreign production, water for chromatography P, solvents for HPLC of Merck company.

Sample preparation:

0.24 g of a crushed medical sponge (powder) was placed in a 25 ml volumetric flask, 15.0 ml of methanol P was added and kept in an ultrasonic bath at room temperature for 30 min, brought the volume of the solution with the same solvent to the mark. The suspension was centrifuged at 5 000 rpm for 10 min, the supernatant was filtered through 0.2 ^m filters before being introduced into the chromatographic system.

Results and discussion:

Was carried out the identification of chlorhexidine digluconate according to the absolute retention time (2.76 min), the corresponding chromatogram is shown on Figure 1.

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Chlorhexidine digluconate, which was prepared by diluting solutions at: 0,8; 2,0; 4,0; 12,0; 20,0 mg/ml. Was observed the linearity of the calibration graph in interval of the concentrations of 0.8-20.0 mg/ml, the lower limit of determination of chlorhexidine digluconate by HPLC was 0.8 mg/ml (Fig. 2).

The chromatogram's of the sample solution of chlorhexidine digluconate in the terms of the quantification of chlorhexidine digluconate in the hemostatic sponges

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Fig. 2: The linear relationship of the area on chlorhexidine digluconate concentration

The regression coefficients were calculated by the least-squares method of the using calibration graph S = bx + a, the correlation coefficient (R) was equal 0,99956.

To determine the quantitative content of chlorhex-idine digluconate in the sponge were prepared test solutions within the studied concentrations. The results of the quantitative determination of the studied AFI in the medical sponge shown in table 1.

Table 1.

The results of determining the quantitative content ^ of chlorhexidine digluconate in the medical sponge

Weight of the piece of the powdered sponge, g The content of chlorhexidine, % Statistical processing of the results of quantitative determination of chlorhexidine digluconate (n = 5; P = 0,95)

0,255 1,02 = 1,02 S2 = 1,85 •lO-4 Sr = 0,0133 S = 0,0136 ■^77 = 0, 0061 ^77 = 0,0168

0,248 1,05

0,261 0,99

0,252 1,03 = 1,65 %

0,257 1,01

Thus, can be used the HPLC method to identify and quantify chlorhexidine digluconate in a sponge, which is a simple, fast, sensitive method of analysis.

CONCLUSION:

1. Were selected the chromatographic conditions and the HPLC method for identification and quantification of chlorhexidine digluconate in the developed medical/hemostatic sponges for stop bleeding or to close wound surfaces of different etiology.

2. Were analyzed the experimental specimens of medical/hemostatic sponge by the developed method; the content of chlorhexidine digluconate was within the defined requirements.

References

1. Hu-Fan Song, Ai-Zheng Chen, Shi-Bin Wang, Yong-Qiang Kang, et al. Preparation of Chi-tosan-Based Hemostatic Sponges by Supercritical Fluid Technology. Materials. 2014; 7: 2459-2473.

2. Vons B.V., Chubka M.B., Hroshovyi T.A. [Tekhnolohichni aspekty rozrobky hubok hemo-statychnykh]. Modern Pharmacy: History, Realities and Prospects for Development: Proceedings of a Scientific Conference with International Participation. 2019;1: 107-108. Ukrainian.

3. Deng A.H., Chen A.Z., Wang S.B. et al. Porous nanostructured poly-l-lactide scaffolds prepared by phase inversion using supercritical CO2 as a nonsolvent in the presence of ammonium bicarbonate particles. J. Supercrit. Fluids. 2013; 77: 110-116.

4. Vons Bohdana, Tryhubchak Oksana, Grochovuy Taras, Chubka Mariana, Bihunyak Vo-lodymyr Research of powders of the cryolyophilized xenoderm of porcine skin. International Journal of Green Pharmacy (IJGP). 2018; 12 (3): 657-664.

5. Jia-Ying Liu, Yang Li, Yang Hu et al. He-mostatic porous sponges of cross-linked hyaluronic acid/cationized dextran by one self-foaming process. Materials Science and Engineering. 2018; 83(1): 160-168. Available from: https://doi.org/10.1016Zj.msec.2017.10.007

6. Wei Sun, Yinghui Chen, Weien Yuan He-mostatic absorbable gelatin sponge loaded with 5-fluorouracil for treatment of tumors. International Journal of Nanomedicine. 2013; 8: 1499-1506.

7. Cardoso M.A., Favero M.L., Gasparetto J.C., et al. Development and validation of an rp-Hplc method for the determination of chlorhexidine and p-Chloroaniline in various pharmaceutical formulations. Journal of Liquid Chromatography & Related Technologies. 2011; 34(15): 1556-1567. Available from: doi:10.1080/10826076.2011.575979.

8. European Parmacopoeia. Strasburg Cedex. 4 ed.: Council of Europe; 2002.

9. Hanin V.A. [Development of methodology for quantitative analysis of active substances of vaginal suppositories "Flugedin"]. Ukrayins'kyy zhurnal klin-ichnoyi ta laboratornoyi medytsyny. 2010; 5(3): 170173. Ukrainian.

10. Davtyan L.L., Voronkina A.S., Chubenko

0.V. et al. Development of methods for quality control of dental medicinal films with chlorhexidine, metronidazole and glucosamine. Farmatsevtychnyy zhurnal. 2016; 3-4: 92-99. Ukrainian.

11. Malkova T.L., Chekrishkin L.A., Evich N.I. Quantitative determination of chlorhexidine biglu-conate in solutions. Farmatsiya. 1998; 3: 37-38. Russian.

12. Berezina E.S., Golovanenko A.L., Alekseeva

1.V. Validation of the quantitative determination

method of chlorhexidine bigluconate in gel for the treatment of dentine caries. Mezhdunarodnyy nauchno-issledovatel'skiy zhurnal. 2017; 5 (58): 133-136. Russian.

13. Sampiiev A.M., Davitavian N.A., Nikiforova Ye.B., Yakuba Yu.F. Quantitative determination of 0.05 % chlorhexidine solution by capillary electrophoresis. Zaporozhye medical journal. 2019; 21(4): P. 517521.

PHARMACOECONOMIC ANALYSIS OF TREATMENT SCHEMES OF OSTEOCHONDROPATHY

IN A HOSPITAL

Stechyshyn I.

PhD (Pharmacology), Department of Pharmacy Management, Economics and Technology I. Horbachevsky Ternopil National Medical University, Ternopil, Ukraine

Kravchuk I.

Student of the Faculty of Pharmacy I. Horbachevsky Ternopil National Medical University, Ternopil, Ukraine

Pavliuk B.

PhD student, Department of Pharmacy Management, Economics and Technology, I. Horbachevsky Ternopil National Medical University, Ternopil, Ukraine

ABSTRACT

Among the many childhood diseases of the musculoskeletal system, a separate group is osteochondropathy, which includes Schinz's (Sever's) disease, which is common among children aged 7-14 years and among athletes. The lack of consensus on the causes the feasibility of modern methods of diagnosis and treatment osteoporosis, this pushes to find a means of influencing the course and prevention of this disease. The purpose of the study was to determine the cost of medical technologies used in the treatment of patients with Schinz disease in hospital. For this purpose, the analysis of the total cost of the disease was used, which allows to take into account the economic aspects when introducing new diagnostic and medical technologies or updated protocols of treatment of certain nosological forms in healthcare institutions.

As a result of the conducted research, it was found that on this basis it is expedient to put into practice the optimal programs of diagnostics, treatment and prevention, which are optimal for their efficiency and cost-effectiveness and which will save society money.

Keywords: osteochondropathy, Schinz (Sever's) disease, heel pain

According to the World Health Organization, diseases of the musculoskeletal system as the cause of disability and mortality rank 4th in the world after cardiovascular, cancer and diabetes [1], and in the near future experts predict an epidemic of osteoporosis, which indicates the aging of the planet's population [2]. According to statistics, every fifth inhabitant of the globe suffers from back pain, and the proportion of osteochondrosis is up to 90%.

Until recently, the opinion was that musculoskeletal disorders are usually characteristic of the elderly, but epidemiological studies, indicate that these phenomena also found among the young people. For example, among healthy children and adolescents of Finland musculoskeletal disorders occur in 20%, Sweden -29%, Switzerland - 51%, Canada - 33% [3].

In Ukraine, chronic diseases of the musculoskeletal system is also one of the most common problems [4]. Thus, according to the results of the State Statistics Service of Ukraine in the Ternopil region only, there are 19096 children with diseases of the musculoskeletal system, which is more than 17% registered among the 89038 healthy children aged 7-14 years. Among the

many diseases of the musculoskeletal system, a separate group is osteochondropathies, which develop in children and adolescents, debuting with pain syndrome or orthopedic disorders that lead to disability over time.

As known from the literature, the incidence of osteochondropathy of the calcaneo-achilles apophysis (Shinz/Sever's disease), a common pathological disease in children and adolescents, which is 3.7 per 1000 registered patients [5]. The actual incidence in the world may be even higher. Unfortunately, there is no data on this disease in Ukraine.

Shinz's disease (Sever's disease) is characterised by pain experienced near the lower posterior aspect of the calcaneus in close proximity to the attachment of the Achilles tendon into the secondary growth plate of the calcaneus. Described in the writings of such scholars as: P. Haglund in 1907, James Warren Sever in 1912 and H. Schinz in 1922. In the territory of the post-Soviet countries, the disease named after the scientist Schinz, and in the world known as a Sever disease [6]. Believed that the disease results from osteopenia, small multiple fractures of the calcaneus, which due to anatomical location are under conditions of excessive mechanical needs [7], disturbance of local blood supply