Analysis of the pharmacological efficacy of methylprednisolone in targeted delivery using biodegradable carriers in animals

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Introduction

The development of novel routes of drug administration is often associated with a number of potential challenges and risks, most are solved through technological im- provements, drug stability studies, and preclinical research [1–3]. Anti-inflammatory drugs for veterinary use are widely represented on the pharmaceutical market. The most accessible options, both in terms of convenience for animal owners and economic considerations, are conventional oral formulations [4–7]. Drugs intended for intravenous administration are preferably given in a veterinary clinical setting to minimize the risks of phlebitis, intravenous contamination, patient injury, and stress. However, beyond these obvious problems, there are also issues related to the direct pharmacological action of drugs and strategies to reduce the drug burden on the animal organism. Targeted delivery using modern biodegradable carriers applied to the surface of implants helps address these issues during the provision of high-tech medical care [8– 10]. The use of targeted drug delivery is promising because it allows the administration of antibiotics, hormones, and anti-inflammatory agents while monitoring their concentrations, reducing drug loss during passage through tissue barriers, and acting directly at the site of implantation injury [9, 11–13]. The use of glucocorticosteroids (GCS) is justified for reducing inflam-matory edema, decreasing granulation of injured tissue, managing allergic reactions, and achieving immunosuppression, all of which affect the rate and effectiveness of implant integration in the animal body [14–15].

The aim of the study is to perform a pharmacokinetic analysis and evaluate the efficacy of methylprednisolone in targeted delivery.

Materials and Methods

For the quantitative determination of GCS concentrations in serum and blood plasma of laboratory animals (rabbits of the Soviet Chinchilla breed, age 5 months, males), four groups of animals (n = 3 males per group) were formed. The first and second groups received methylprednisolone hemisuccinate in the dosage form of a solution for intramuscular and intravenous injections; the third and fourth groups received a methylprednisolone-based formulation. Rabbits in groups 1 and 2 received methylprednisolone hemisuccinate intravenously at a dose of 20.0 mg/kg, while ani- mals in groups 3 and 4 received an equivalent dose intramuscularly. Megestrol acetate was chosen as an internal standard for the analytical study. The standard sample was added directly to the test samples during sample preparation. The use of a standard  sample during sample preparation serves as a calibration mechanism, allowing the determination of minimal concentrations of the target GCS in the serum and plasma of the experimental animals.

Results and Discussion

The concentrations of methylprednisolone hemisuccinate and methylpred- nisolone were studied in in vivo experiments. Biological material was collected from animals at the same time (10:00 a. m.) after standard procedures. The sample volume was 2.0 mL; blood was collected into Eppendorf tubes. The analysis was performed in two media — plasma and blood serum — to determine differences in drug concentrations and metabolism in the body. The results are presented in Tables 1, 2 and Figs. 1–4.

Table 1
The content of GCS in the plasma and serum of rabbits after intravenous administration of a dose of 20.0 mg/kg

№	Time	Methylprednisolone hemisuccinate, mcg/ml (No. 1)		Methylprednisolone, mcg/ml (No. 3)
		Plasma	Serum	Plasma	Serum
1	0 min	0.0	0.0	0.0	0.0
2	15 min	12.9	11.5	0.7	0.4
3	30 min	5.3	5.1	1.1	0.9
4	2 hours	2.7	2.0	2.1	1.9
5	4 hours	2.3	2.1	2.1	2.2
6	24 hours	0.2	0.2	0.0	0.0

Source: compiled by A.N. Ibragimova, J. Bannoud, S. Mas-Koma, N.I. Troshina, A.S. Karamyan.

Table 2
The content of GCS in the plasma and serum of rabbits after intramuscular administration of a dose of 20.0 mg/kg

№	Time	Methylprednisolone hemisuccinate, mcg/ml (No. 2)		Methylprednisolone, mcg/ml (No. 4)
		Plasma	Serum	Plasma	Serum
1	0 min	0.0	0.0	0.0	0.0
2	15 min	3.6	3.4	0.4	0.2
3	30 min	2.9	2.8	0.7	0.5
4	2 hours	1.6	1.3	1.7	1.4
5	4 hours	0.4	0.3	2.0	2.1

Source: compiled by A.N. Ibragimova, J. Bannoud, S. Mas-Koma, N.I. Troshina, A.S. Karamyan.

During the quantitative determination of GCS concentrations in the plasma and serum of the experimental animals, the limit of detection for methylprednisolone hemisucci- nate was established at 20 pg/mL in all animals. At the same time, the concentration of methylprednisolone was twice that of methylprednisolone hemisuccinate.

Fig. 1. Dependence of the concentrations of methylprednisolone hemisuccinate and methylprednisolone in the serum of rabbits (intravenous administration of a dose of 20.0 mg/kg)
Source: compiled by A.N. Ibragimova, J. Bannoud, S. Mas-Koma, N.I. Troshina, A.S. Karamyan.

Fig. 2. Dependence of the concentrations of methylprednisolone hemisuccinate and methylprednisolone in the plasma of rabbits (intravenous administration of a dose of 20.0 mg/kg)
Source: compiled by A.N. Ibragimova, J. Bannoud, S. Mas-Koma, N.I. Troshina, A.S. Karamyan.

Fig. 3. Dependence of the concentrations of methylprednisolone hemisuccinate and methylprednisolone in the plasma of rabbits (intramuscular administration of a dose of 20.0 mg/kg)
Source: compiled by A.N. Ibragimova, J. Bannoud, S. Mas-Koma, N.I. Troshina, A.S. Karamyan.

Fig. 4. Dependence of concentrations of methylprednisolone hemisuccinate and methylprednisolone in the serum of rabbits (intramuscular administration of a dose of 20.0 mg/kg)
Source: compiled by A.N. Ibragimova, J. Bannoud, S. Mas-Koma, N.I. Troshina, A.S. Karamyan.

To confirm the stability of the drug on the surface of the biodegradable implant carrier, samples were analyzed using a well-established and standardized method of high-per- formance liquid chromatography with tandem mass spectrometry (HPLC–MS/MS). The concentration of methylprednisolone was determined by studying the finished carriers. The conducted release allowed us to establish the drug concentration on the film in the first week at 723 ± 18 μg. A repeat analysis of the samples after two weeks revealed a decrease in the drug content on the film, with a loss of 3.59% (less than 5%), which is acceptable (Fig. 5).

Fig. 5. Graph of the release of methylprednisolone from microcells on biodegradable carriers in phosphate buffer
Source: compiled by A.N. Ibragimova, J. Bannoud, S. Mas-Koma, N.I. Troshina, A.S. Karamyan.

Conclusion

A comparative in vivo study of GCS concentrations in the plasma and serum of experimental animals established significantly higher values in plasma. The correlation between the release, biotransformation, and elimination of methylprednisolone depending on the proposed route of administration was investigated. The demonstrated release of GCS from the film applied to the implant supports the expanded use of biodegradable surfaces and the improvement of pharmacotherapeutic approaches using glucocorticos- teroids without increasing postoperative risks.

About the authors

Georges Bannoud

Vetlife Veterinary Clinic

Email: george911@mail.ru
Veterinarian 12 Beskudnikovsky Blvd., bldg. 1, Moscow, 127474, Russian Federation

Alfiya N. Ibragimova

RUDN University

Email: Ibragimova-an@rudn.ru
ORCID iD: 0000-0003-3484-3949
SPIN-code: 3948-5218

Candidate of Pharmaceutical Sciences (PhD), Associate Professor, Department of Disaster Medicine, Medical Institute

6 Miklukho-Maklaya St., Moscow, 117198, Russian Federation

Santiago Mas-Coma

University of Valencia

Email: S.Mas.Coma@uv.es
ORCID iD: 0000-0002-1685-7004

Doctor of Veterinary Sciences, Professor, Director of the Department of Veterinary and Animal Science

Av. de Vicent Andrés Estellés s/n 46100, Burjassot, Valencia, Spain

Natalya I. Troshina

RUDN University

Email: troshina-ni@rudn.ru
ORCID iD: 0009-0003-8230-0153
SPIN-code: 9355-7573

Senior Lecturer, Department of Veterinary Medicine, Agrarian and Technological Institute

6 Miklukho-Maklaya St., Moscow, 117198, Russian Federation

Arfenya S. Karamyan

RUDN University

Author for correspondence.
Email: karamyan-as@rudn.ru
ORCID iD: 0000-0003-2112-673X
SPIN-code: 5511-4446

Doctor of Biological Sciences, Candidate of Veterinary Sciences (PhD), Associate Professor, Associate Professor of the Department of Veterinary Medicine, Agrarian and Technological Institute

6 Miklukho-Maklaya St., Moscow, 117198, Russian Federation

References

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