INTRODUCTION:

Some diseases, such as ulcerative colitis, can affect only the lower part of the gastrointestinal tract (large intestine). These disorders may need the topical action of the active agent at the inflamed parts, where a topical drug dosage form may not be effective alone in treating some severe and specific cases.¹Most orally delivered drugs are absorbed in the upper part of the gastrointestinal tract and such systems may fail to effectively treat large-intestinal inflammatory diseases.²This advocates the necessity to develop a large-intestinal specific oral drug delivery system, capable to deliver the active moiety to specific parts of the large intestine to enhance the therapeutic efficacy, decrease needed dose and reduce potential side effects. For this purpose, an ideal oral delivery system should be capable to retain the active moiety until reaching the large intestine, where it acts topically at the inflamed parts.³

Any orally delivered agent is estimated to stay a maximum of 8 hours in the upper part of the gastrointestinal tract; this include 1 - 4 hours in the stomach and 3 ± 1 hours in the small intestine.⁴ This has paved the way to design a drug delivery system that is capable to retard the drug release during the first 6 hours (following its oral administration) and permit its release afterward in a controlled way. This can ensure the drug delivery to the lower part of the gastrointestinal tract (large intestine), where the residence time is estimated to be around 10 hours.⁵

Mesalazine, also known as mesalamine, is the first choice cure in treating large-intestinal inflammatory diseases, where it is believed to exert local relief effect at the affected inflamed parts. It is usually prescribed as a topical preparation for mild cases and as an oral preparation in addition to the topical one in severe cases.⁶A previous study reported the development of a delayed delivery system based on long hydrophobic chains, as potential system to deliver drugs to the large intestine. Ibuprofen and mesalazine were used in this system as model drugs, the in vitro release evaluations had showed the ability of this system to achieve a relatively higher drug release in the simulated large intestinal fluid compared to that in the simulated gastric and small intestinal fluids.⁷In another work, a mesalazine loaded nanocarrier, based on polyurethane-grafted chitosan, was developed as a delayed drug delivery system. The release studies had demonstrated the potential applicability of this system to delay the mesalazine release, with no burst effect and less than 60% of the loaded drug was released after 8 hours.⁸In another study, mesalazine tablets were produced using different concentrations of hydroxypropyl methyl cellulose and ethyl cellulose as release retarding polymers. The optimized formulation exhibited around 86 % drug release after 6 hours in the simulated intestinal fluid and it was concluded that it can be promising as a colon specific delivery system.⁹

Human made monomers have been widely used to synthesize crosslinked polymers in many pharmaceutical and biomedical applications. They are usually chosen over the natural ones due to many reasons; such as the ability to produce large uniform quantities, reproducible characteristics and the flexibility to modify the copolymerized structure.¹⁰Hydroxyethyl methacrylate is the first and most famous used synthetic monomer in the pharmaceutical industry and it has been reported to be biocompatible, chemically as well as thermally stable.^11-14A previous study reported the use hydroxyethyl methacrylate monomer to synthesize a cationic crosslinked polymer with dimethylaminoethyl methacrylate monomer, using poly ethylene glycol diacrylate as a crosslinker, for the potential delivery of anti-cancer drug. Doxorubicin was used as a model drug and in vitro release studies were performed in buffers of pH 7.4 and pH 5.5 to simulate healthy and cancer site respectively. A four times fold drug release was obtained at pH 5.5 compared to that at pH 7.4, which indicated the potential applicability of this carrier as an anti-cancer delivery system.¹⁵ Another study has also employed hydroxyethyl methacrylate monomer in the synthesis of an anionic polymer, designed for the effective and safe oral delivery of nonsteroidal anti-inflammatory therapeutic agents. The produced formulations were loaded with ketoprofen as a model drug. A higher in vitro release and swelling was obtained in simulated intestinal fluid compared to that in simulated gastric fluid, which indicated the potential success of this system.¹⁶

The main aim of our study was to develop a crosslinked poly 2-hydroxyethyl methacrylate carrier loaded with mesalazine as a model drug, synthesized by bulk polymerization method, using different crosslinker concentrations, as a promising large-intestinal drug delivery system. Swelling and in vitro release evaluations were conducted for the produced carriers.

MATERIALS AND METHODS:

Materials:

Mesalazine, ethylene glycol dimethacrylate (EGDMA), hydroxyethyl methacrylate (HEMA), sodium chloride, disodium hydrogen phosphate dodecahydrate, azobisisobutyronitrile (AIBN), potassium chloride, potassium dihydrogen phosphate, sodium hydroxide and sodium dodecyl sulphate were purchased from Sigma-Aldrich. HPLC grade water used in all experiments. All materials were used as purchased with no modifications.

Methods:

Preparation of polymeric carriers using different concentrations of EGDMA:

The HEMA based polymeric formulations were synthesized successfully by bulk polymerization technique using the recipes shown in Table 1. A mold for each formula was initially prepared using two pyrex glass sheets (214 x 200 X 5mm), silicon release liner, silicone medical tubing and three foldback clips (32 mm). Firstly, the two pyrex sheets were placed horizontally on the bench and the liner sheet sited above them. after that, the tubing was used to define the mold's borders and finally the clips were used to hold the pyrex sheets together in a vertical way. The ingredients of each recipe were mixed together at room temperature with stirring for 1 hour, then infused into the predesigned mold using an injection syringe and finally putted in a preheated oven (60˚C) to initiate the synthesis process, which lasted for18 hours at 60˚C. The produced polymeric formulations were place in separated storage boxes and soaked with HPLC grade water, changed every day, to wash the produced formulations and eliminate any unreacted or unwanted species. This stage lasted for a week and a UV/VIS spectrophotometer (Spectroscan 80 D, Biotech Engineering Ltd., UK) was used to assure its end. All swollen formulations were pierced using a cork borer (6.25 mm) to produce uniform pellets, which were dried until achieving a constant weight.

Table 1: The synthesized polymeric formulations

Formula	HEMA (% w/w)	EGDMA (% w/w)	AIBN (% w/w)	Mesalazine (% w/w)
C1	98	1	1	-
C2	94	5	1	-
C3	89	10	1	-
M1	96	1	1	2
M2	92	5	1	2
M3	87	10	1	2

Swelling studies:

The swelling behavior of the produced carrier pellets was investigated at 37°C in a swelling medium of pH 7.4 buffer. Each pellet was initially weighed, then immersed in a 6ml buffer, previously maintained at 37 °C, inside a glass vial. The vials were then putted inside a static water bath at 37°C. At specific time points (1/12, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 24, 48, 72, 96 and 120 hours), forceps were used to withdraw the immersed pellets and place them on a thick tissue paper to remove pellets surface liquid. After that, the pellets were weighed and immersed again in the same medium inside the water bath. Five replicates of each formulation were carried out. Equation 1 was used to find the swelling ratio obtained at each specific time point. A plot of the obtained selling ratios versus the specific time points (hours) was done for the pellets to study the swelling behavior and investigate the effect of EGDMA concentration. A GraphPad Prism 8 software was used to analyze all data statistically using a two way analysis of variance, followed by Tukey’s multiple comparisons test (n=5, p < 0.05). The swelling data of the first 10 hours (prior reaching the plateau) were also fitted to the Korsmeyer Peppas model to estimate the swelling rate of the prepared pellets.

The swollen pellet weight- The dried pellet weight

Swelling ratio (%) = -------------------------------------------- X 100 ---(1)

The swollen pellet weight

In vitro release studies:

The release profile of mesalazine from the produced pellets was investigated, using a modified method of Heelan and Corrigan, in pH 7.4 buffer with 1% (w/v) SDS at 37°C.¹⁷Forceps were used to dip the pellets in a 20ml release medium, previously kept at 37°C, inside a wide mouth glass bottle sealed with aluminum screw cap. The release study was performed in a shaking bath operating at 100 round per minute and 37°C. A sample of 0.5ml was withdrawn from each bottle, and replaced with the same volume of fresh medium (previously kept 37°C) at specific time points (5, 60, 120, 180, 240, 300, 360, 420, 480, 540, 600, 660, 720 and 1440 minutes). The withdrawn samples were then filtered, and the mesalazine release was quantified using a UV/VIS spectrophotometer (Spectroscan 80 D, Biotech Engineering Ltd., UK) via measuring the absorbance at 330 nm. The concentration of mesalazine was then calculated using a fully validated calibration curve of mesalazine at 330nm. Three replicates of each pellet were carried out. The release profile of mesalazine from each formulation was obtained by plotting the cumulative release ratios versus the specific time points (minutes). A GraphPad Prism 8 software was used for the statistical analysis of the release data using a two way analysis of variance, followed by Tukey’s multiple comparisons test (n=3, p < 0.05). The first 60% of all release data were fitted to the Korsmeyer Peppas model to find the release rate constants and investigate the release mechanism.¹⁸

RESULTS AND DISCUSSION:

Preparation of polymeric carriers using different concentrations of EGDMA:

As described in Table 1, polymerized formulations based on hydroxyethyl methacrylate were synthesized successfully by a free radical polymerization method and using different concentrations of EGDMA, as a crosslinker (1, 5 and 10% w/w). The main objective of producing these polymerized pellets was to develop a time dependent drug delivery system, which is capable to retard the drug release within the first 6 hours of its administration and control its release afterward; to deliver the drug mainly to the large intestine. Mesalazine was used as a model drug because it is commonly used to treat inflammatory disorders associated with the colon.¹⁹

Swelling studies:

The ability of a polymeric material to swell in a fluid is usually governed by the material–fluid and material-material interactions.²⁰The maximum swelling is achieved when a balance occurs between these two interactions.²¹ The swelling behaviour of the produced polymeric pellets was investigated in pH 7.4 buffer using a previously described quantitative method. Figures 1-3 show the effect of EGDMA concentration on the swelling behaviour of the produced pellets (C1-C3). The equilibrium swelling ratio was achieved within 24 hours for the three carriers with 33%, 27% and 20% ratios obtained for C1, C2 and C3 respectively. Increasing the crosslinker concentration has significantly reduced the swelling capacity of the produced pellets. This can be contributed to the fact that increasing the crosslinker concentration results in a more rigid structure and reduces the polymer elasticity.^22,23 The estimated swelling rate constants (Table 2) have also confirmed this result; as increasing EGDMA concentration decreased the swelling rate constant of the produced pellets (0.18, 0.16 and 0.13 for C1, C2 and C3 respectively). An equation was obtained following the linear regression of the equilibrium selling ratios versus the crosslinker concentration (Figure 3). This equation can give a prediction of the maximum swelling capacity that can be attained in pH 7.4 buffer upon using a specific EGDMA concentration and vice versa.

Figure 1: Time-dependent swelling ratio (mean ± SD, n=5) of C1, C2 and C3 in PBS.

Figure 2: Equilibrium swelling ratio (mean ± SD, n=5) of C1, C2 and C3 in PBS.

Figure 3: Simple linear regression of equilibrium swelling ratio (mean ± SD, n=5) vs EGDMA Conc. (% w/w).

Table 2: Swelling rate constants and R² after fitting the swelling data to the Korsmeyer Peppas model

Formulation	R²	Swelling rate constant
C1	0.986	0.176
C2	0.990	0.160
C3	0.997	0.128

In vitro release studies:

The in vitro release profiles of the produced pellets are shown in Figure 4. A delayed mesalazine release was achieved in the three produced pellets and they demonstrated the ability to retard the drug release within the first 6 hours; as around 34%, 24% and 16% cumulative drug release were obtained from M1, M2 and M3 respectively. The three pellets have also exhibited the ability to control mesalazine release up to 24 hours, where 76%, 62% and 41% were obtained for M1, M2 and M3 respectively. The ability to retard the drug within the first 6 hours and control it afterward indicates the potential applicability of the produced pellets as successful large-intestinal delivery systems. Increasing the crosslinker concentration had significantly decreased the drug release profile and this agree with the swelling studies. This can be explained by the formation of a more rigid polymeric network and reduced polymer elasticity.^22,23 The EGDMA concertation effect on the release profile was also confirmed by the release rate constants in Table 3; as 0.12, 0.08 and 0.05 were obtained for M1, M2 and M3 respectively. Three mechanisms (diffusion, swelling and chemically controlled) are usually believed to, separately or collectively, control drug release from polymerized drug delivery systems.^24-26 The Korsmeyer Peppas release model relates the fraction of drug released with time via a simple exponential equation (Equation 2), and it is frequently used to investigate the release mechanisms in polymerized carriers.^27,28In equation 2, M refers to the fraction of drug released at a specific time point (t, hours), Symbol K refers to the release rate constant and n refers to the release exponent that is used to indicate the release mechanism. These parameters can be estimated after the linear regression of log M versus log t, where the slope represents n and K is the antilog of Y intercept. The interpretation of the release mechanism depends on the estimated n value. A Fickian diffusion is interpreted when this value is less than or equal 0.5, An anomalous mechanism is indicated when it is greater than 0.5 and less than 1.0, case 2 transport when the value equals 1 and super case 2 transport when the value is greater than 1.^29,30 As shown in Table 3, the n value was greater than 0.5 and less than 1 for all pellets. This indicates an anomalous release, which means the drug release is controlled by diffusion and the carrier swelling.

Figure 4: The release profiles of mesalazine from the prepared pellets (mean ± SD, n=3) at pH 7.4.

M = K t^{n
………………………..}(2)

Table 3: R², n and K_m after fitting the release data to the Korsmeyer Peppas model

Formulation	R²	n	K
M1	0.959	0.581	0.117
M2	0.951	0.605	0.083
M3	0.933	0.622	0.052

CONCLUSION:

A free radical polymerization technique was used to successfully synthesize polymeric drug carriers, based on hydroxyethyl methacrylate monomer and using different crosslinker concentrations. Mesalazine was used in these carriers as a model drug. The developed carriers demonstrated their ability to delay the in vitro release of the model within the first 6 hours; as around 34%, 24% and 16% cumulative drug release were obtained from M1, M2 and M3 respectively. Additionally, theses carriers exhibited a controlled drug release within 24 hours with cumulative drug release of 76%, 62% and 41% obtained for M1, M2 and M3 respectively. Using a higher concentration of the crosslinker had significantly decreased the swelling and release profiles of the produced carriers, which was explained by the formation a more rigid polymeric network with reduced elasticity. The swelling and in vitro release studies had proven the ability of the developed pellets to delay the drug release during the first 6 hours of their administration, and control the release up to 24 hours, which makes them promising as large-intestinal drug delivery systems.

ACKNOWLEDGEMENTS:

The authors are grateful to the Philadelphia University, Amman, Jordan for the research support (No. 511/34/100 PU).

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Received on 02.12.2020 Modified on 16.05.2021

Research J. Pharm. and Tech 2022; 15(1):35-39.

DOI: 10.52711/0974-360X.2022.00007