(T1330-08-54) Do the pH-Stage Dissolution Tests Lie? Case Study of Duloxetine Hydrochloride Dissolution in the Small-Scale Setup

Purpose: Biorelevant in vitro simulation of gastrointestinal (GI) tract environment in dissolution tests is a crucial step in formulation development and predicting of a drug performance in clinical trials, where extended number of tests are often necessary, especially for pH-sensitive chemicals and modified-release formulations e.g. resistance to gastric environment. Standard dissolution tests require substantial resources. Reducing the scale test from 1000 mL to just 25 mL can increase the number of performed tests without consuming a significant amount of valuable Active Pharmaceutical Ingredients (APIs). The aim of this study was to design a small-scale approach for dissolution testing under biorelevant conditions including simulation of gastric and intestinal compartments. We have investigated the influence of: a) pH of gastric conditions, b) the buffering system simulating intestinal fluids and c) pH-stage or gradual change of pH on the dissolution profile of gastro-resistant formulation.

Methods: Duloxetine Zentiva 60 mg hard capsules (Zentiva, k.s., Czech Republic), containing duloxetine hydrochloride in the form of gastro-resistant pellets with hydroxypropylmethylcellulose acetate succinate (HPMCAS) coating, were used as a model drug formulation. Dissolution tests were conducted using the in-house designed small-scale dissolution setup. Online monitoring of pH was provided with pHysiograd® (Physiolution GmbH, Germany). The volume of the single sample was set to 25 mL and the temperature was maintained at 37 ºC. The pH-stage experiments started in simulated gastric fluid (SGF) at pH 1.2, 2.0 or 4.0 (purified water, adjusted with hydrochloric acid). After 30 min, relevant Fasted State Simulated Intestinal Fluid (FaSSIF, Biorelevant.com Ltd, UK) concentrate was added to simulate gastric emptying (GE) and change into the intestinal environment. The pH-stage experiments were conducted for 120 min. In the first row of tests, the FaSSIF concentrate based on phosphate buffer was added and the pH was manually adjusted to 6.5. In the second row of tests, the intestinal conditions were provided with FaSSIF based on a bicarbonate buffer, where pH at 6.5 was automatically adjusted with pHysiograd® using gaseous titrants. The last test was designed to simulate real GI conditions. In the gastric stage pellets were incubated in hydrochloric acid at pH 2.0 for 30 min. The intestinal pH gradient was conducted in FaSSIF based on a bicarbonate buffer. The pH was gradually increased from 5.81 to 7.69 in approx. 220 min of the experiment to reflect pH gradients recorded in a group of patients [1]. All experiments were conducted in triplicate (n=3). The duloxetine hydrochloride concentration was measured using a modified HPLC method adapted from Patel, et al. [2].

Results: The results were calculated as the amount of released duloxetine dose (%) over time. We did not record the API in the gastric conditions. In the pH-stage experiments, pellets started to disintegrate between 10 to 20 min after the change of pH to 6.5, i.e. 40-50 min since the beginning of the test. We observed the impact of the first medium on the dissolution performance of duloxetine in simulated intestinal fluids. We noticed the lowest levels of released API in FaSSIF over 120 min of the experiments in the tests started at pH 1.2 under gastric conditions, while significant differences between the impact of SGF at pH 2.0 and 4.0 were not recorded. We noticed a higher amount of duloxetine released in FaSSIF based on the phosphate buffer than in the bicarbonate buffer-based for all tested conditions. In the pH-gradient experiment, we recorded API to start dissolving after 60 min since the beginning of the test, but the significant increase in released duloxetine was observed after 105 min. The concentrations similar to those observed in 120 min of pH-stage experiments were recorded by us after 180 to 240 min of the pH-gradient test. This is related to the pH-dependent disintegration of HPMCAS coating which is triggered at pH > 5, however when the pH is lower than 6.0 the disintegration process lasts more than 60 min, while at pH 6.5 the time is approx. 20 min depending on the grade of the polymer [3,4].

Conclusion: Conducting dissolution tests on a small scale (resource-saving approach) including pH-stage experiments, as well as dynamic pH-changes is feasible. The pH of the simulated gastric fluid had an impact on subsequent pellets disintegration - physiologically unlikely conditions of pH 1.2 affected the polymer coating slowing down its disintegration. The pH-stage experiments do not reflect physiological conditions i.e., biorelevant GI pH-gradients and their changes over time. This may cause misinterpretation of overestimated performance. Implementation of a bicarbonate buffer in dissolution tests enables dynamic pH adjustments and more biorelevant simulation of GI conditions on a small scale without changing the volume and ionic strength of the medium.

References: 1. Koziolek M., Grimm M., Becker D., et al. Investigation of pH and Temperature Profiles in the GI Tract of Fasted Human Subjects Using the Intellicap® System, J. Pharm. Sci., 2015; 104 (9), 2855-2863, doi:10.1002/jps.24274.
2. Patel SK, Patel NJ, Patel KM, Patel PU, Patel BH. Estimation of Duloxetine Hydrochloride in Pharmaceutical Formulations by RP-HPLC Method. Indian J Pharm Sci. 2008; 70(6), 825-827. doi:10.4103/0250-474x.49136.
3. Brady J., Dürig T., Lee P.I., Li J.-X.. Chapter 7 - Polymer Properties and Characterization,
Editor(s): Qiu Y., Chen Y., Zhang G. G.Z., Yu L., Mantri R.V. Developing Solid Oral Dosage Forms (Second Edition), Academic Press, 2017; 181-223, doi:10.1016/B978-0-12-802447-8.00007-8.
4. Sarabu S., Kallakunta V.R., Bandari S., et al. Hypromellose acetate succinate based amorphous solid dispersions via hot melt extrusion: Effect of drug physicochemical properties. Carbohydr Polym., 2020, 1;233:115828. doi: 10.1016/j.carbpol.2020.115828.

Acknowledgements:

Funding: none
Disclaimer: This study does not require ethics committee approval for animal use or human participation.

Figure 1. Duloxetine release profiles obtained using the small-scale setup. Data expressed as a percentage of dose (MW+SD, n=3).