Formulation and Pharmaceutical Assessment of Annona muricata Oral Capsules and Suspension as Antidiarrhea Dosage Forms

Department of Pharmaceutics, Faculty of Pharmacy and Pharmaceutical Sciences, Kwame Nkrumah University of Science and Technology, Kumasi, Ghana Department of Herbal Medicine, Faculty of Pharmacy and Pharmaceutical Sciences, Kwame Nkrumah University of Science and Technology, Kumasi, Ghana Department of Pharmaceutics, School of Pharmacy, Central University, Miotso, Ghana Department of Pharmaceutics and Microbiology, School of Pharmacy, University of Ghana, Accra, Ghana


Introduction
Diarrhea is a condition of having unusual loose or watery stools with a sense of urgency. It is also known to occur when the lining of the intestine is unable to retain fluid, leading to lose stool. e precarious nature of the disease is linked with dehydration due to fluid loss and consistently high morbidity in the absence of vigorous interventions. Diarrhea is a leading cause of death in all age groups with an estimated 1.31 million deaths recorded annually across the globe for a mortality rate of 7.58% [1,2]. e burden of deaths is greatest in low-income countries such as Ghana where getting access to portable water and proper sanitation is a problem. In Ghana, the mortality rate as at 2020 was 7.15% per 1000 inhabitants with diarrhea accounting for about 25% of death in children under five years with correspondingly high debilitating effects in adults [1][2][3][4].
Most patients with diarrhea do not seek medical attention unless symptoms last for more than three days or they have coexisting fever, prostration, or rectal bleeding by which time their condition may have worsened. Hence, it is very imperative that medications used in the treatment of diarrhea are effective and readily available [1,3]. In order to treat diarrhea, various orthodox medicines have been developed. However, the World Health Organization promotes studies on medicinal plants that can be used to manage and treat diarrhea and to formulate these medicinal plant extracts in standardized dosage forms that will be convenient, efficient, and safe for the patient. In Ghana, traditionally, a lot of plants are used to treat diarrhea including Bridelia ferruginea, Albizia ferruginea, and Psidium guajava (guava). Another plant commonly used to treat diarrhea in Ghana is Annona muricata [5][6][7].
Annona muricata of the family Annonaceae, popularly known as soursop, graviola, or guanabana is a fruit that grows in the tropical regions of America, Africa, and the Caribbean. Phytochemical studies carried out on various parts of the plant revealed the presence of tannins, phenolic compounds, flavonoids, and alkaloids which underpin their pharmacological relevance [6,8]. Further research carried out by Doe et al. shows that the ethanolic extract of the seeds have antidiarrheal property and corroborates its folkloric use [9]. Irrespective of the reported dose and pharmacological activity of the ethanolic extract of the seeds of Annona muricata in the management of diarrhea, its use is limited in clinical settings due to the absence of standardized dosage forms. Against this background, this study seeks to provide standardized dosage forms (suspensions and capsules) of the ethanolic extract of the seeds of Annona muricata in order to substantiate its clinical usefulness and integration into current health care delivery as an effective antidiarrheal agent.

Collection of Sample and Extractive Procedure.
Annona muricata fruits were obtained from the Ashaiman market, in the Greater Accra region of Ghana, and authenticated at the Department of Pharmacognosy, KNUST, Kumasi, Ghana. Extraction was done using a previously established method [9]. e seeds were removed from the fruits. It was then dried under shade for 7 days, and the weight was taken afterwards. e dried seeds were milled into fine powder and weighed as well. e milled seeds were then macerated in 80% ethanol in an airtight container and stored in the refrigerator at 5°C for 7 days. e decanted solution was filtered using a filter paper, and the filtrate was evaporated using a rotary evaporator and then freeze dried into fine powder.

Assessment of the Flow Characteristics.
e indirect method described by Aulton and Taylor [10] in the characterization of the flow properties of materials was used.

Formulation of Annona muricata Suspension.
In the formulation of 100 mL suspension, Annona muricata extract was weighed and added to the previously weighed Acacia and triturated with benzoic acid and double strength chloroform water to obtain a uniform mixture (such that each 5 mL contains 0.15 g). Water was then used to top up to the mark (Table 1). e excipients were selected after several preformulation studies with different excipients.

Formulation of Annona muricata Granules.
In preparing the granules, starch, lactose, and Annona muricata extract were weighed and mixed using geometric dilution (Table 2). e powdered mixture was then blended with tragacanth mucilage to form a damp mass and prepared into granules (14.8 g) using the method described by [10]. e excipients were selected after several preformulation studies with different excipients.

Evaluation of Flow Properties of Annona muricata
Granules.
e flow properties of the formulated granules were determined using the fixed height method for angle of repose and bulk density measurements (Hausner ratio and Carr's index) [10].

Encapsulation of Annona muricata Granules.
e granules were encapsulated using capsule size 1 with 0.15 g of extract in each capsule (150 mg per dose). Fifty-five (55) capsules were filled simultaneously.

Quality Assessment of Formulated Capsules.
e weight uniformity and disintegration tests were determined as described by the British and International Pharmacopeias [11,12].
2.7.1. Drug Content. Ten capsules were used for this experiment. A capsule was emptied, and 20 mL of 0.1 M HCl was added. e mixture was filtered, and 0.1 M HCl was used to top it up to 50 ml. Drawell's UV spectrometer was used in measuring the absorbances at 300 nm after serially diluting the solutions. e recorded absorbance was then inserted into a calibration equation, and the percentage content was determined for each capsule [11,[13][14][15].

In Vitro Dissolution Test.
In vitro dissolution study was done using the USP type I basket dissolution apparatus. 900 mL of 0.1 M HCl was transferred into each of the six dissolution vessels, the temperature of the dissolution vessel was maintained at 37 ± 2°C, and a randomly selected capsule was dropped into the dissolution medium. An aliquot of the sample was withdrawn at regular time intervals (5,15,30,35,40,45, and 60 minutes), and the same volume of 0.1 M HCl was used to replace the volume withdrawn. e samples were filtered and analyzed with the UV spectrometer at 300 nm. e absorbances obtained were inserted into the calibration equation to obtain the amount of drug released at each time point. A graph of cumulative drug released against time was then plotted to obtain the dissolution profile of the formulated capsules in 0.1 M HCl [11, 13-15].

Uniformity in Drug Content of Formulated
Annona muricata Suspension. Five (5) mL of the formulated suspension was diluted with 25 mL of 0.1 M HCl, filtered, and topped up to 50 mL using the same solvent (0.1 M HCl). Drawell's UV spectrometer was used in measuring the absorbances at 300 nm. e recorded absorbance was then inserted into the calibration equation, and the percentage content was determined. is was repeated for nine other 5 mL aliquots [11,[16][17][18].

In Vitro Dissolution Study of Annona muricata Suspension.
e USP type II paddle dissolution apparatus was used with 900 mL of 0.1 M HCl (as the dissolution medium) heated to 37°± 2°C. e paddles were set to a speed of 50 rpm, and 5 ml of the prepared suspension was transferred into the dissolution medium. After 5, 15, 30, 35, 40, 45, and 60 minutes, an amount of 10 mL of the dissolution medium was withdrawn, filtered, and the absorbance at 300 nm was determined using Drawell's UV spectrometer. e withdrawn volume was replaced with same volume of 0.1 M HCl to maintain sink conditions. e absorbances obtained were inserted into the previously determined calibration equation to obtain the amount of drug released at each time point. A graph of cumulative drug released against time was then plotted to obtain the dissolution profile of the formulated suspension in 0.1 M HCl [11,[16][17][18]].

Flow Characteristics of Granules and Extract.
e powdered extract showed good flow properties with Hausnerʼs ratios of 1.12, Carr's index of 10.98%, and angle of repose of 20.5°, while the Annona muricata granules exhibited excellent flow properties with Hausnerʼs ratios of 1.10, Carr's index of 8.93%, and angle of repose of 17.5°. is confirms that the formulated granules will improve the encapsulation process by enhancing the uniform filling of the capsule shells [11,13].

Capsule Weight Uniformity.
e weight uniformity of Annona muricata capsules as given in Table 3 complied with the weight uniformity specifications according to the International Pharmacopeia [12].
is implies that each manufactured capsule potentially has the exact amount of the Annona muricata extract and excipients specified, with little variability, reducing the risk of under dosing and overdosing.

Disintegration Profile of Formulated Capsules.
Based on the results given in Table 4, the formulated capsules passed the disintegration test as stipulated by the British Pharmacopeia [11].
is indicates that the capsules will release their contents within time for further dissolution of the extract to occur.

Drug Content.
e percentage drug content of all ten sampled capsules were within the standard range of 85.0-115% ( Table 5) and confirms that the encapsulation process as well as other preencapsulation procedures were carried out accurately.
e results also indicate that the formulated capsules will provide the required therapeutic dose without any overdose or underdosage that would be harmful or detrimental to the patient.

In Vitro Dissolution of Annona muricata Extract from
Capsules. According to the data (Figure 1), 99.29% of the extract was released at the 45th minute. is implies that the capsules passed the dissolution test and can dissolve well in physiological solution, allowing the extract to be absorbed and the intended pharmacological activity to be achieved [11,13].

Appearance Test.
Over the course of four weeks, the formulated suspension was closely monitored for the appearance or absence of undesired physical changes. ere were no unwanted physical changes in the prepared suspension (Table 6). is shows that the suspension is physically stable and retains its aesthetic attractiveness and that the excipients employed in the formulation were appropriate [10,19].

Sedimentation Volume and Rate of Formulated Suspension.
A good suspension, in order to improve uniform and accurate dosing, should not have the suspended particles settling quickly. e sedimentation volume of an ideal suspension should be close to 1 with a correspondingly low sedimentation rate [19][20][21]. is property was evident in the formulated suspension ( Figure 2).

Rheology, Redispersibility, and pH of Formulated
Suspension. ere was no statistical variation (P ≥ 0.05) in the pH of the formulated suspension (Table 7) to indicate any physicochemical change [19,22,23]. e flow time of the suspension was short with a correspondingly high apparent viscosity for easy pourability (Table 7) [ 16,19,22]. roughout the study period, the solid particles were redispersed with less than ten (10) strokes (Table 7). e prepared suspension possessed pseudoplastic flow, which implies that it may be swiftly redispersed with modest agitation, and the appropriate amount can be easily drawn (Figure 3). is feature is critical for a good suspension [24].

Drug Content.
All sampled doses passed the drug content test (Table 8) [11]. is implies that each volume will have the exact amount of extract needed, preventing overdosing and underdosing, both of which are hazardous to the patient's health.

In Vitro Dissolution Profile of Annona muricata Extract from Formulated Suspension.
According to the data (Figure 4), 99.20% of the extract was released at the 45th minute which complies with pharmacopeia specifications. is implies that the capsules can dissolve well in physiological solution, allowing the extract to be absorbed and eliciting the intended therapeutic effect [11,22].      Week Physical instability (aggregation, caking, crystal growth formation) 1 No physical instability observed 2 No physical instability observed 3 No physical instability observed 4 No physical instability observed

Conclusion
is study has shown that the ethanolic extract of Annona muricata can be successfully formulated into capsules and suspension. e prepared dosage forms satisfied the pharmacopeia requirements for quality evaluation and may be used as appropriate alternatives in the management and treatment of diarrhea.

Data Availability
e data used to support the findings of this study are included within the article and also available from the corresponding author upon request.

Conflicts of Interest
e authors declare that there are no conflicts of interest.