Release Formulations for Certain Newer drugs
Dissertation submitted to
The Tamilnadu Dr. M.G.R. Medical University, Chennai, in partial fulfillment of the requirements
for the award of the Degree of
DOCTOR OF PHILOSOPHY
K.KRISHNARAJ, M.Pharm.,
Under the Guidance of Dr. M.J.N. CHANDRASEKAR
J.S.S. College of Pharmacy, Rocklands, Ootacamund 643 001,
India.
J.S.S. COLLEGE OF PHARMACY, ROCKLANDS, OOTACAMUND – 1
DECLARATION
I hereby declare that the thesis entitled “Design and development of controlled/sustained release formulations for certain newer drugs” submitted by Mr. K.Krishnaraj to The Tamilnadu Dr.M.G.R. Medical University, Chennai, in partial fulfillment of the requirements for the award of Degree of Doctor of Philosophy in Pharmaceutical Sciences, is the result of his original and independent research work carried out at J.S.S. College of Pharmacy, Ootacamund, during the years 2007-2010, under my supervision. The thesis or any part thereof has not formed the basis for the award of any degree, diploma, associateship, fellowship, or any other similar title, of this or any other University, previously.
(Dr.M.J.N.CHANDRASEKAR) Supervisor
OOTACAMUND - 1
DECLARATION
I hereby declare that the thesis entitled “Design and development of controlled/sustained release formulations for certain newer drugs” submitted by Mr.K. Krishnaraj to The Tamilnadu Dr.M.G.R. Medical University, Chennai, in partial fulfillment of the requirements for the award of Degree of Doctor of Philosophy in Pharmaceutical Sciences, is the result of his original and independent research work carried out at J.S.S. College of Pharmacy, Ootacamund, during the years 2007-2010, under the supervision of Dr. M.J.N. Chandrasekar, Professor, J.S.S. College of Pharmacy, Ootacamund. The thesis or any part thereof has not formed the basis for the award of any degree, diploma, associateship, fellowship, or any other similar title, of this or any other University, previously.
OOTACAMUND - 1
DECLARATION
I hereby declare that the thesis entitled “Design and development of controlled/sustained release formulations for certain newer drugs” submitted by Mr.K. Krishnaraj to The Tamilnadu Dr.M.G.R. Medical University, Chennai, in partial fulfillment of the requirements for the award of Degree of Doctor of Philosophy in Pharmaceutical Sciences, is the result of his original and independent research work carried out at J.S.S. College of Pharmacy, Ootacamund, during the years 2007-2010, under the supervision of Dr. M.J.N. Chandrasekar, Professor, J.S.S. College of Pharmacy, Ootacamund. The thesis or any part thereof has not formed the basis for the award of any degree, diploma, associateship, fellowship, or any other similar title, of this or any other University, previously.
(Dr.M.J.NANJAN) Director
I hereby declare that the thesis entitled “Design and development of controlled/sustained release formulations for certain newer drugs” submitted by me to The Tamilnadu Dr. M.G.R. Medical University, Chennai, in partial fulfillment of the requirements for the award of Degree of Doctor of Philosophy in Pharmaceutical Sciences, is the result of my original and independent research work carried out at J.S.S. College of Pharmacy, Ootacamund, during the years 2007-2010, under the supervision of Dr. M.J.N. Chandrasekar, Professor, J.S.S. College of Pharmacy, Ootacamund. The thesis or any part thereof has not formed the basis for the award of any degree, diploma, associateship, fellowship, or any other similar title, of this or any other University, previously.
(K. KRISHNARAJ) J.S.S. College of Pharmacy
S.No Description Page No
1 Introduction 1
Literature review 18
2 Scope and objectives of the present Study 28
Proposed plan of work 32
Plant profiles 35
3 Experimental 39
Materials 39
Isolation and physicochemical characterization of the
polysaccharides 39
In vivo toxicity studies of the polysaccharide 46
Preformulation studies 47
Development of sustained release tablets 49
Stability studies of the developed formulations 53
Development of sustained release spheroids 53
Bioanalytical method development and validation 56
Bioavailability studies 62
4 Results And Discussion 67
Purification and characterization of the polysaccharides 67
In vivo toxicity studies of the GG 69
Preformulation studies 70
Drug content and physical evaluation 72
In vitro dissolution studies of QF 74
In vitro dissolution studies of OH 77
Stability studies 79
Formulation of spheroids of QF by extrusion spheronization 79
Validation of HPLC method 82
Bioavailability studies of the selected drugs in healthy rabbits 86
Human bioavailability studies 86
5 Summary and Conclusions 89
6 References 92
1
1.
Introduction
This thesis deals with the investigations carried out by the writer on the development of controlled / sustained drug delivery systems using a novel polysaccharide isolated from Delonix regia for quetiapine fumarate (QF) and ondansetron hydrochloride (OH). Before discussing the experimental procedures adopted and the results obtained, a brief introduction to sustained drug delivery systems including oral sustained release drug delivery systems, followed by site-specific delivery schemes to the gastrointestinal tract (GI tract) would be presented here. Based on the mechanism of drug release, the different polymeric drug delivery systems including targeted drug delivery systems will also be briefly discussed. The use of the selected drugs, the problems associated with their use, a literature survey on the investigations that have been carried out by earlier investigators to isolation of polysaccharide from plant sources along with the earlier work controlled/sustained release formulations on the drugs selected if any would also be presented.
1.1. Historical perspective
patient-2 compliant dosage forms. A significant increase has been noted in approvals of newer drug delivery systems in the past couple of years and this is expected to continue at an impressive rate in the future1.
1.2. Oral sustained release
Sustained release (SR) technology has rapidly emerged over the past three decades as a new interdisciplinary science that offers novel approaches to the delivery of bioactive agents into systemic circulation at a predetermined rate. The choice of the drug to be delivered, clinical needs, and drug pharmacokinetics are some of the important considerations in the development of SR formulations, in addition to the relationship between the rates of drug release from the delivery system to the maximum achievable rate of drug absorption into the systemic circulation. By developing a predictable and reproducible drug release rate for an extended period of time, SR formulations can achieve optimum therapeutic responses, prolonged efficacy and also decreased toxicity2. The therapeutic advantages of SR systems over conventional dosage forms have been amply documented in the literature3, 4. One of the important advantages is the reduced dosing frequency, which improves patient compliance and therapeutic efficacy, in addition, to constant blood levels of the drug, unlike in conventional systems as shown in Figure 1.
Figure 1. Plasma drug concentration vs. time profile
3 Oral administration is the most popular route due to ease of ingestion, pain avoidance, versatility (to accommodate various types of drug candidates) and most importantly, patient compliance5-7. Also, solid oral delivery systems do not require sterile conditions and are, therefore, less expensive to manufacture.
1.3. Currently marketed oral sustained release approaches
Advances in oral sustained release technology are attributed to the development of novel biocompatible polymers and machineries that allow preparation of novel design dosage forms in a reproducible manner. The main oral drug delivery approaches that have survived are, coating technology using various polymers for coating tablets, nonpareil sugar beads and granules matrix systems made of swellable or non-swellable polymers, slowly eroding devices and osmotically controlled/sustained devices
1.4. Sites of action for sustained release
4 1.5. Polymeric drug delivery systems
A strong relationship exists between polymer chemistry and drug delivery systems. Other than mechanical pumps, all drug delivery systems depend on polymeric materials. From its modest beginning in 1973 when drugs were simply mixed with polymeric matrices to control their release, tremendous advances have been made during the past decades on polymeric drug delivery systems.8,9 These advances that have been made are fueled not only by a convincing need for these systems but also by powerful commercial interests. Significant contributions have been made by researchers associated with industrial research programmes in close collaboration with the scientists in the academic field. The article by Sinko and Kohn gives an excellent overview of polymeric drug delivery systems10.
1.6. Classification of polymeric drug release systems
Drug delivery systems can be broadly divided into two large categories, namely, sustained drug release systems and targeted drug release systems, based on the relationship between the site of drug release and the site of drug action. Unlike in the case of sustained drug release systems where the drug is delivered into the systemic circulation at a predetermined rate, targeted drug release systems deliver the drug predominantly to the site of action. An advantage of targeted drug delivery is that high local concentrations of the drug can be achieved since the drug is delivered predominantly to the site of action rather than being distributed throughout the whole body.
5 classified as ‘matrix systems’ and ‘reservoir systems’.11 In the matrix system the drug is uniformly dispersed within the polymer, whereas in the reservoir system the drug is in a separate phase and physically dispersed within a surrounding and rate limiting polymeric phase. Both matrix and reservoir systems can be formulated in a wide range of sizes and shapes, from micro particles to large discs and slabs.
Based on the mechanism of drug release, polymeric drug release systems can be classified as solvent controlled systems, diffusion controlled systems and chemically controlled systems. Though this classification is based on theoretical considerations, in practical situations the rate of drug release is affected by various combinations of the above three mechanisms.
1.6.1. Solvent controlled systems
Solvent controlled systems can be further divided into swelling controlled systems and osmotic pumps. Swelling controlled systems are generally hydrogels, for example polymers that have been made insoluble by cross linking. In these systems the rate of swelling and hence the rate of drug release depends on the hydrophilic and hydrophobic balance between the polymeric matrix and the degree of cross linking12.Several hydrogel matrices that swell or shrink in response to temperature and pH changes have also been investigated.13-15 A number of other factors can be used to control the release of drugs from polymers16.
6 1.6.2. Diffusion controlled systems
Reservoir and matrix systems are the two diffusion controlled drug release systems. Reservoir systems have been formulated as capsules, microcapsules, hollow fibers or tubes with sealed ends. Here the rate of drug release is strongly sustained by the rate of the diffusion of the drug through the polymeric membrane. Two different types of membranes are generally used, namely homogenous membranes and microporous membranes. In microporous membranes, as in Transderm Scop system that uses a polypropylene membrane, the drug diffuses through the pores that are filled with the same medium as the reservoir, whereas in homogenous membranes, as in Transderm-Nitro system that uses an EVA copolymer, the rate of diffusion of the drug depends on the partitioning between the membrane and the drug16. A disadvantage with the implantable drug reservoir system is the danger of ‘dose dumping’. The polymeric membrane sometimes becomes leaky due to cracks as a result of which the entire drug core is released to the circulation within a short time. Both nondegradable and degradable membranes have been used depending on the requirement of the patient, the ease with which the system can be removed and the general medical environment. Polyanhydride based drug release system for BCNU is an example of biodegradable matrix systems and Nitro-Dur and Nitro-Dur II transdermal systems are based on nondegrdable matrix systems.17, 18
1.6.3. Chemically controlled systems
Systems in which the rate of drug release is predominantly controlled by the rate at which the drug is cleaved from the polymer backbone or the rate at which the polymer degrades, the rate of physical erosion of the polymer, are called chemically sustained systems. The two main types of chemically controlled systems are, matrix systems based on degradable polymers and pendant chain systems based on soluble macromolecular drug conjugates.
7 device is slower than the rate at which the polymer disintegrates into water soluble substances. In other words, the transformation of the polymer into water soluble substances takes place only at the outer surface of the solid device.19-21.
1.6.4. Targeted drug delivery systems
The basic premise of a targeted drug delivery system is the assumption that the therapeutic index of a drug can be improved when the drug accumulates selectively in specific tissues or organs or cell types. The essential characteristics of an ideal targeting system have been defined by Mills and Davis22 as follows;
• Compatibility with the body in terms of toxicity, biodegradability and antigenicity,
• Protection of the drug until it reaches its site of action,
• Maintenance of the drug-carrier integrity until the target is reached, • Avoidance of interaction with normal cells,
• An ability to travel intervening membranes, • Target recognition and association,
• Sustained drug release to achieve the desired therapeutic effect and • Carrier elimination from the body following the drug release.
8 of the accepted strategies to avoid RES uptake and target to sites other than the RES (liver, spleen, etc) is to prepare colloidal systems less than 50-60 nm so as to enable them for transendothelial passage via fenestrations or sisusoids28.
Unlike in the case of low molecular weight drugs, soluble polymeric drug conjugates are internalized into cellular compartments by a process called pinocytosis. Pinocytosis is the only process by which most water soluble polymeric drug conjugates enter cellular compartments and what is more pinocytosis occurs in almost all cell types. The polymeric drug conjugates that are internalized by the pinocytic process ultimately accumulate in the lysosomal compartments of the cell which has an acidic pH environment depending on the cell type. The hydrolytic enzymes present in the lysosomal compartments are capable of cleaving and releasing the drugs from their polymeric carriers. The bond between the polymeric carrier and the drug should of course be susceptible to enzyme catalyzed hydrolysis and not affected during transport in the blood streams (plasma and serum) and the extracellular species. An impermeable drug can thus be sent inside the cellular compartments by conjugating it with a suitable water soluble polymeric carrier. This type of drug delivery is referred to as lysosomotropic drug delivery and was established by DeDuve in 197429. Duncan and Kopechek 30,31 have eventually investigated on this type of drug delivery systems.
1.7. Site-specific delivery in the GI tract
In recent years there has been a dramatic increase in both the number and the rate at which new potential drug candidates are being moved into clinical trials. One of the initial screening criteria for these new drug candidates is their oral bioavailability. Many of the potential and exciting drug candidates are passed over because of their poor or variable absorption profiles. Site-specific delivery in the GI tract may provide a solution to some of these otherwise poorly absorbed drug candidates 32, 33.
9 biochemical issues have to be taken into consideration. Some of these have been described in detail by Mrsny32.
The most direct and simplest method of site-specific delivery along the GI tract is that of a direct drug application like a simple mouth wash to the oral cavity or an enema to the rectum34. Enema formulations have been shown to spread and coat the entire descending and rigmoid colonic segments as well as the rectum. Suppository deliveries can provide essentially the same site-specific delivery advantages as enema delivery. Though patients sometimes eliminate suppositories before their complete dissolution, there are definite advantages to rectal delivery that includes avoiding first-pass metabolism and reducing the mucosal metabolism in some cases.
Site-specific delivery of drugs can also be made through chemical conjugates, polymeric carriers and devices. Such approaches take advantage of the unique environments present in the GI tract that occur as a result of normal physiological events or pathological status. Devices have also been developed to deliver drugs to different segments of the GI tract. Polymeric drug reservoirs retained at the buccal mucosa using bioadhesive polymers have shown promise35, 36. Polysulfone microporous hollow fibers have been used as platform to deliver drugs to the buccal cavity37.
10 1.8. Hydrophilic matrices
Hydrophilic matrix tablets are among the most popular delivery systems for oral controlled/sustained release dosage forms. These hydrophilic matrices are widely accepted because of their biopharmaceutical and pharmacokinetic advantages over conventional dosage forms38. This is largely because they offer precise modulation of drug release as a result of hydration of the constituent polymer(s), flexibility to obtain the desired drug release profiles, cost effectiveness, patient compliance, providing a constant, prolonged, and uniform therapeutic effect and broad Food and Drug Administration (FDA) acceptability39. From the wide choice of possible matrix materials like sodium alginate, chitosan, poly (acrylic acids), etc., hydroxypropyl methylcellulose (HPMC) has been used most frequently in the formulation of sustained release monolithic matrix tablets because of its hydrophilic gel-forming property, non-toxicity and cost effectiveness40–42. The swelling rate and erosion of HPMC-based matrix tablet in aqueous media are very crucial in terms of achieving the desired release profiles, and are affected by parameters such as the physicochemical properties of the polymer and the drug, processing conditions, the testing medium used and the formulation composition43–45.
Hydrophilic matrix systems have been used extensively to produce sustained drug delivery through the gastrointestinal (GI) route. These gels have been shown to produce near zero order drug release kinetics46-48. Such matrix formulations often contain HPMC, although other polymeric materials have also been evaluated49,50. The operating principle for controlling the drug release from hydrogel matrix tablets on exposure to aqueous fluids is well known and has been shown to be a complex interaction between swelling, diffusion and erosion51.
11 capable of being processed into tablets using either direct compression, following the addition of drug and lubricant powder, or conventional wet granulation. Particulate variations, for instance, following filling into a hard gelatin capsule, have also been produced, tested and shown to retain sustained release properties. Drug release from tablets containing the polysaccharide excipient system has been found to be capable of control using a variety of different formulations and process methods. This provides a variety of different release modalities that are capable of matrix dimension independence, as well as independence from influence from the bulk properties of granules produced. Advantages over sustained release systems composed of single components of the system can also be demonstrated. The hydrogel excipient system can be used in a conventional wet granulation process if desired.52 Although a variety of dosage forms have been developed for the preparation of oral SR formulations, they broadly fall into two categories: single unit dosage forms and multiple (multiparticulate) dosage forms.
1.9. Single unit dosage forms
Single unit dosage forms are defined as oral dosage forms that consist of single units, with each unit containing one dose of the drug and intended to be administered singularly. There are several such dosage forms that have been developed for the sustained release of various bioactive materials, as has been reported in the literature of which monolithic matrix-based tablets are the most common single unit dosage form used for sustained drug delivery53,54. Advantages associated with such dosage forms include high drug loading, simple and cost-effective manufacturing, the availability of a wide range of excipients and polymers for controlling the drug release and the possibility of using different mechanisms for controlling the drug release such as diffusion controlled, swelling controlled, erosion sustained or a combination of all of these. Single unit dosage forms that have been used for sustained drug delivery include drug-release controlling polymer, membrane-coated tablets and osmogen-sustainedformulations55, 56.
12 lubricants and other excipients. Using a suitable rate controlling polymer, the matrix can be tabletted by direct compression or conventional wet granulation method. Matrix systems are highly resistant to release inconsistencies and dose dumping since they are relatively simple systems and are robust to process and ingredient variations, production methods and end use conditions. A hydrophilic matrix sustained release system is easy to formulate and easy to produce. It is a robust dynamic system composed of polymer wetting, hydration and dissolution. Water penetrates into the tablet, causing the gel layer to become thicker. The soluble drug diffuses out of the gel layer at a rate sustained by the gel viscosity. With soluble drugs, the primary release is by diffusion through the gel layer. With insoluble drugs, the primary mechanism is by the tablet surface erosion. The drawback of matrix-type delivery systems is their first-order drug delivery mechanism caused by changing surface area and drug diffusional path length with time57.
1.10. Polysaccharide based hydrophilic matrices
The principles of oral sustained release medication and their performance and mechanisms of the action of hydrophilic sustained release matrices have been reviewed extensively elsewhere 58-60 A wide variety of different hydrogel materials have been described for use in sustained release medicines. Some of these are synthetic61,62 but most are of semi-synthetic or natural origin63-66 and relatively few contain both synthetic and non-synthetic material.67 However, some of the systems require special processes and production equipment. In addition, some are susceptible to variable drug release as the result of one or more of the following effects: pH dependency68; food effect variability69; ions and ionic strength dependency; viscosity dependency; corrosion/erosion variability; content uniformity problems; flow and weight uniformity problems; carrying capacity and mechanical strength problems. Additionally, formulation of a specific hydrophilic matrix-forming system to provide other than a first order drug delivery regime has often been problematical.
13 contact with water and these have been used for the preparation of single unit dosage forms73-75. The powdered drug is embedded uniformly in a matrix of the hydrogel and compressed to form a tablet, a production method that is relatively simple and cheap to perform. They find a wide range of pharmaceutical applications that include their use as binders and disintegrants in tablets, emulsifiers, suspending agents, and gelling agents. They are also used as sustaining agents in tablets. Many of these gums and mucilages have been reported to sustain the drug release from matrix tablets76,77. Tablets are among the simplest drug delivery systems and hence the obvious choice for oral delivery of drugs.
1.11. Multiple unit dosage forms
The concept of the multiple unit dosage form was initially introduced in the early 1950s. These forms play a major role in the design of solid dosage form processes because of their unique properties and the flexibility found in their manufacture. These forms can be defined as oral dosage forms consisting of a multiplicity of small discrete units, each exhibiting some desired characteristics. Together, these characteristic units provide the overall desired sustained release of the dose. These multiple units are also referred to as pellets, spherical granules or spheroids.
1.12. Pellets, spherical granules or spheroids
14 Compaction, drug layering, melt spheronization, globulation, balling, compression, and extrusion spheronization are the methods that have been reported for the preparation of pellets. Among these, extrusion-spheronization is the most popular method in pharmaceutical industries85. Although the main thrust of spheronization processes is to manufacture spherical drug cores that will be subsequently coated to modify the drug release, it is also possible to prepare spheroid cores that inherently possess specific release profiles. This is achieved by the incorporation of substances that modify the drug release. These substances are called release modifiers. To retard the drug release, polymers such as shellac and waxes are used86.
Pellets should possess sufficient strength to overcome any appreciable abrasion during agitation.
The strength of the pellets depends, to a great extent on the physical forces that bound the primary particles together. Although, initially mechanical forces such as tumbling, kneading, agitation, extruding, rolling and compression are needed to bring individual particles in contact with one another.
1.12.1 Challenges in pellet formulations
• Dose adjustment, content uniformity different size and large dose.
• Due to low density of pellets may float on the surface of GI and high density would be in the bottom or under the surface.
• Dose adjustment can be done using placebo pellets mixing but reproducibility cannot be obtained.
• They may not reach the therapeutic level for pellets containing potent drugs if missed in one pellet too.
1.12.3. Methods of pellet preparation
15 mechanism might also exist that is based on frictional forces as well as rotational forces. In this mechanism a twisting of the cylinder occurs after the formation of a cylinder with rounded edges, finally resulting in the breaking of the cylinder into two distinct parts with both parts featuring a round and a flat side. The edges of the flat side fold together like a flower, forming the cavity observed in certain pellets because of the rotational and the frictional forces involved in the spheronization process. Figure 2 shows both pellet-forming mechanisms.
Figure 2. Pellet forming mechanisms.
(a)Rowe: I Cylinder; II. Cylinder with rounded edges; III.Dumb-bell; IV. Ellipse; V.Sphere
(b)Baert : I Cylinder; II.Rope; III. Dumb-bell; IV.Sphere with a cavity outside; V. Sphere
16 • The first approach involves the placement of a drug in an insoluble matrix in which the eluting medium penetrates the matrix and the drug diffuses out of the matrix and into the surrounding pool for ultimate absorption.
• The second approach involves enclosing the drug particles with a polymer coat. In this case, the portion of the drug that has been dissolved in the polymer coat diffuses through an unstirred film of liquid into the surrounding fluid.
• The third approach is eroding beads in which the drug is released as the bead matrix erodes or dissolves.
i) Advantages of the extrusion and spheronization process • Ease of operation
• High throughput with low wastage • Narrower particle size distribution • Production of pellets with low friability
• Production of pellets are suited for film coating
• More sustained and better sustained drug-release profile when compared with other techniques
ii) Process and equipment
• Granulation : preparation of the wet mass • Extrusion : shaping the wet mass into cylinders
• Spheronization : breaking up the extrudate and rounding off the particles into spheres
• Drying : drying of the pellets.
iii) Mechanism of drug release from sustained release pellets
• The placement of a drug in an insoluble matrix in which the eluting medium penetrates the matrix and the drug diffuses out of the matrix and into the surrounding pool for ultimate absorption
• Enclosing the drug particles with a polymer coat. In this case, the portion of the drug that has been dissolved in the polymer coat diffuses through an unstirred film of liquid into the surrounding fluid
17 1.12.4. Coated pellets
Sustained drug release from pellets is conventionally achieved by polymer coating. The active ingredients releases according to zero-order kinetics.
1.12.5. Matrix pellets
Sustained release from pellets is conventionally achieved by polymeric coating. There is growing interest in the development of matrix pellet formulations because, in practice, polymeric coating is associated with various problems like.
• The process is time consuming and expensive. • Film thickness is variable.
• There may be cracks in the film or aging of the polymer coating.
• The drug release profile is not reproducible because of inconsistent film coating and
• Coating is dependent on the optimization of several parameters during the production process.
Three classes of release-retarding materials are used for the formulation of matrix SR formulations. They include insoluble or ‘skeleton’ matrices, water-insoluble but erodible matrices and hydrophilic matrices. Examples of the materials used as release retardants are the followings;
i) Insoluble and inert
Polyethylene, polyvinyl chloride, methyl acrylate-methacrylate copolymer, ethyl cellulose
ii) Insoluble erodable
Carnauba wax, stearyl alcohol, stearic acid, polyethylene glycol, polyethylene glycol monosterate, triglycerides
iii) Hydrophilic matrices
18 1.13. Literature Review
A review of the literature was carried on the, isolation of polysaccharides from plant sources and their use in matrix tablets and spheroids along with sustained release formulations if any on the selected drug candidates.
Investigation of polysaccharides from plant exudates and Drumstick and Tamarind
Subhas and coworkers90 have carried out investigations on the purification of the whole gum exudate from the drum-stick plant (Morirzga oleifera) and showed that L-arabinose, D-galactose, u-glucuronic acid. L-rhamnose, D-mannose, and D-xylose were present in the gum in the molar ratio of 14.5: 11.3: 3: 2: 1: 1. A homogeneous, degraded-gum polysaccharide consisting of galactose, glucuronic acid and D-mannose in the molar ratio of 11.7: 3.9: 1 was obtained on mild hydrolysis of the whole gum with acid. Per-methylation studies were conducted on the whole gum, the degraded gum, and their carboxyl-reduced products. The results were in good agreement with those obtained from periodate oxidation. Isolation and characterization of the oligosaccharides obtained from the mother liquor during the preparation of the degraded gum and by graded hydrolysis of the degraded gum were also achieved. On the basis of the results obtained from these studies a tentative structure was assigned to the average repeating unit of the gum.
Panda and coworkers91 have investigated the gelling potential of a natural gum obtained from Moringa oleifera exudates. The gum was extracted by using water and precipitated using acetone as non solvent. Physical charecterstics such as solubility, swelling index, loss on drying and pH were studied. Diclofenac sodium was used as a model drug for formulating gels. Seven batches of drug loaded gels with concentration of mucilage ranging from 5.5, 6.0, 7.0, 7.5, 8.0 and 8.5 were formulated by using glycerine as a plasticizer and methyl paraben as the preservative. The gels prepared with 8.0% of mucilage were found to be ideal and comparable with commercial gels.
19 precipitation. The phytochemical screening and physiochemical properties of the isolated polysaccharide proved its suitability for sustained release purposes. Sustained release matrix tablets were prepared using propranalol hydrochloride by the wet granulation method. The binding property of the gum was also investgated using different concentrations of the gum solution as a binder. The in vitro drug release studies with different polymer concentrations and different fillers (Calcium sulphate dihydrate, lactose) showed prolonged drug release from matrix tablets and moderate binding capacity.
Sadhan and coworkers93 have isolated a water soluble polysaccharide from the aqueous extract of the pods of Moringa oleifera. The polysaccharide contains D-galactose, 6-O-Me-D-D-galactose, D-galacturonic acid, L-arabinose, and L-rhamnose in the molar ratio of 1:1:1:1:1. On the basis of total hydrolysis, methylation analysis, periodate oxidation, and NMR studies a tentative structures were assigned.
Durcilene and coworkers94 have isolated a polysaccharide from cashew tree exudes and carboxymethylated the same in aqueous alkaline medium using monochloroacetic acid (MCA) as the etherifying agent. Ian M. Sims and coworkers95 have isolated a gum that exudes from the wounded trunk of the New Zealand native tree Meryta sinclairii.They also carried out a complete chemical investigation and the structure of the gum and compared it with the structure of gum Arabic.
20 Puja Goyal and coworkers97 have investigated a polysaccharide isolated from tamarind kernel powder and showed that it is a rich source of xyloglucan and can be utilized in a number of pharmaceutical industries. With a view to utilize the gum for broader applications, carboxymethylation of the tamarind kernel powder was carried out. The reaction conditions were optimized with respect to concentrations of sodium hydroxide, monochloroacetic acid, solvent ratio, reaction time and the reaction temperature. Carboxymethylation of tamarind kernel powder increased its solubility in cold water and the stability of its paste to microorganisms.
Investigation on Jack fruit and Durian fruit
Amiza and coworkers98 isolated durian seed gum from defatted local durian. The yield was 18% of light brown crude gum powder. Further purification was carried out by barium complexing to give a yield of 1.2% of pure air-dried gum or 0.5% of freeze-dried gum. The pure durian seed gum was characterized in terms of moisture, ash content, mineral content, effect of temperature and pH on viscosity and sugar composition of the gum. The purified durian seed gum had 17.9% moisture and 29.8% total ash. Mineral content of the gum was comparable to commercial gum except for zinc content which was quite high in durian seed gum. The gum solutions were fairly stable over a wide range of pH (2.0–10.0). Sugar analysis by PC and HPLC revealed the presence of L-rhamnose, glucose and D-galactose sugars in the hydrolyzed durian seed gum in the ratio of 3:9:1.
21 Suresh and coworkers100 have isolated an α-D-galactose-specific lectin from the seeds of jack fruit (Artocarpus integra) in pure form by affinity chromatography on immobilized guar gum (a galactomannan). The lectin was shown to be a glycoprotein containing 3% carbohydrate having a molecular weight of 39,500 as determined by gel filtration. Sodium dodecyl sulphate gel electrophoresis revealed a single polypeptide of 10,500 dalton, indicating that the native lectin is a tetramer of identical subunits. The hemagglutinating activity of the lectin towards erythrocytes of all blood groups was found to be the same.
Investgation on Delonix regia and seed gum
Silvana and coworkers101 have isolated a serine proteinase inhibitor from the Delonix regia seeds, a leguminosae tree of the Caesalpinioideae subfamily. The inhibitor, named DrTI, inactivated trypsin and human plasma kallikrein with Ki values 2.19X10-8 M and 5.25 nM, respectively. Its analysis by SDS-PAGE 10–20% showed that the inhibitor is a protein with a single polypeptide chain of Mr 22 h Da. The primary sequence of the inhibitor was determined by Edman degradation that indicated 185 amino acids and showed that it belongs to the Kunitz type family. Its reactive site did not contain Arg or Lys at the putative reactive site (position 63, SbTI numbering).
V. P. Kapoor102 has isolated galacto mannan composed of n-galactose (1 mol) and D-mannose (2 mol) from the seeds of Delonix regia. Hydrolysis of the methylated galactomarkn yielded 2,3,4,6-tetra-O-methyl-o-galactose (l.02 mol), 2.3,~tri-O-methyl-D-mannose (1.05 mol) and 2,3-di-0-2.3,~tri-O-methyl-D-mannose (1 mol). The periodate consumption was l-30 mol for each hexose unit with concomitant liberation of O-31 mol -of formic acid. Hydrolysis of the reduced oxopolysaccharide gave only glycerol (1 mol) and erythritol (1.04 mol). It was established that galactomannan is a highly branched polysaccharide consisting of the main chain of mannose united linked through α (l-4) and the side chain of single galactose units linked through β (1-6).
22 and solved by molecular replacement method and refined, respectively, to R-factor and R-free values of 21.5% and 25.3% respectively at 1.75A resolution. The structure has a classical b-trefoil fold that different from canonical Kunitz type (STI) inhibitors. Its reactive site loop has an insertion of one residue, Glu68, between the residues P1 and P2. Surprisingly DrTI is an effective inhibitor of trypsin and human plasma kallikrein, but not of chymotrypsin and tissue kallikrein. Putative structural grounds of such specificity were discussed.
Brummera and coworkers104 have isolated fenugreek gum, from the defatted and deactivated fenugreek seeds (produced in Canada) at 10.8oC for 2 h with an yield of 22% with only 2.36% protein contaminate. Further purification of the fenugreek gum was achieved by treating the gum solution with pronase to reduce the protein contaminates to 0.57%. Monosaccharide and methylation analysis suggested that the extracted fenugreek galactomannans was highly substituted and the ratios of galactose to mannose were from 1.00:1.02 to 1.00:1.14. Although fenugreek gum exhibited higher molecular weight compared to locust bean gum and guar gum, its the intrinsic viscosity and rheological behavior of fenugreek gum were reduced. This was attributed to the influence of the substitution patterns of the galactose on the mannosyl backbone chain. The purified fenugreek gum demonstrated less surface activity compared to the unpurified gum.
23 Sarika Gupta and coworkers106 have isolated a polysaccharide from the seeds of Cassia occidentalis, an annual weed occurring throughout India, and a rich source of galactomannan gum. The gum derived from seed endosperm can be potentially utilized in a number of pharmaceutical industries to replace the conventional gums. With a view to utilize the gum for broader applications, carbamoylethylation of C. occidentalis seed gum was carried out with acrylamide in presence of sodium hydroxide under different reaction conditions. Variables studied were concentration of sodium hydroxide, acrylamide, gum–solvent ratio, reaction time and temperature. The nitrogen content, carboxyl content and total ether content were also determined.
Vandana and coworkers107 have isolated on a non-ionic water-soluble galactomannan having a galactose and mannose in 1:2 molar ratio from endosperm of the seeds of Ipomoea turpethum. The seed gum was shown to have a branched structure consisting of a linear chain of β(1 -4) linked mannopyranosyl units with D-galactose side chains attached through α(1-6) linkage to the main chain, a fundamental structural pattern found in other seed galactomannans like guar, carob, locust bean, tara and dhaincha commercial gums. The seed gum from Ipomoea turpethum showed similarity in structural pattern and properties to guar gum. I. turpethum seed gum is thus in a natural form and after modification by grafting may find use as a commercial gum.
24 Spheroids
Consuelo and coworkers109 have evaluated the utility of super disintegrants, namely, croscarmellose sodium and sodium starch glycolate in microcrystalline cellulose extrusion–spheronization pellets as a means of increasing the dissolution rate of poorly water-soluble drugs. The model drug used was hydrochlorothiazide, with water or water/ethanol as wetting agent for pellet preparation. Neither disintegrant had any significant effects on pellet morphology, flow properties or mechanical resistance. Nevertheless, the disintegrants afforded a modest increase in drug dissolution rate, attributable to the observed increase in pellet micropore volume. Drug dissolution rate was slightly higher in pellets prepared with sodium starch glycolate, probably because of this disintegrant’s higher swelling capacity.
Steven and coworkers110 have revealed that carbopol 974P NF resin could be incorporated into beads manufactured by extrusion and spheronization and can slow the release of a highly water soluble drug. The complex nature of the extrusion and spheronization process and the various components in the bead formulations, a statistically sound factorial experiment was considered for this study. A one-half fraction of a two level factorial design with three center points was employed to estimate the effects of simultaneously modifying multiple process and formulation variables, including the carbopol concentration, calcium chloride concentration, water content, and the spheronization speed and time. Product yield, average bead roundness, and the drug release profile were selected as responses. Increasing the carbopol content across the experimental range resulted in a significant reduction in the percentage drug released at 25, 40, and 60 min. The results suggest that combining the conditions of high carbopol, high water and low calcium chloride levels with low spheronization speeds at long spheronization times produce the highest quality bead with the prolong drug release.
25 quantities were analysed. The morphological and mechanical characteristics of the obtained beads were also investigated. With demineralized water as the granulation fluid, pellets with a maximum of 50% (m/m) of chitosan could be produced. The mass fraction of chitosan within the pellets could be increased to 100% by using diluted acetic acid for the granulation step.
Diva and coworkers112 have evaluated the effects of certain process variables in the feasibility of producing Microcel MC 101 pellets by the extrusion/spheronization technique. A 23 factorial design was realized to demonstrate the influence of the significant factors and their interactions in the experimental response. The selected process variables such as water content, extruder screen size and spheronizer speed were studied, as well as their influences on the properties of particle size distribution and the densities. The results showed that high levels of the three factors increased sphere size, and low levels decreased it. A strong interaction between water content and extruder screen size was observed for the particle size distribution response. Extruder screen size has a significant effect on the bulk density. Water content and spheronizer speed interaction was shown to influence the sphere density.
Jittima and coworkers113 have prepared pellets by extrusion and spheronization containing microcrystalline cellulose (MCC) and four model drugs with decreasing order of solubility, paracetamol (P), diclofenac sodium (D), ibuprofen (IB) and indomethacin (IN) at a 10% level with and without the addition of a range of levels of glyceryl monostearate (GMS). In spite of these differences in extrusion performance, it was possible to prepare satisfactory pellets from formulations of all the drugs with 0, 30 and 60% GMS combined with 90, 60 or 30% of MCC at a range of water levels. It was also possible to prepare pellets containing the drug D with 70, 80 and 90% GMS, with corresponding quantities of 20, 10 and 0% of MCC. It was also possible to prepare pellet formulations by dispersing the drugs in molten GMS, grinding and processing this with MCC and water.
Ondansetron hydrochloride
26 benzalkonium chloride (BC) and 10% sulfobutylether ß-cyclodextrin sodium salt (SBCD), was somewhat more rapid up to 1.5 h compared to the addition of 10% PEG. The permeation flux increased as the drug concentration increased regardless of the vehicles used. The addition of nicotinamide or chitosan to the aqueous drug solution (40 mg/ml) with 10% PEG 300 and 0.01% BC rather decreased permeation rate and delayed the lag time. Even though cyclodextrins including SBCD or dimethyl-ß-cyclodextrin failed to show permeation enhancing effects of OH, they showed the addition of 10% SBCD to an aqueous solution containing 10% PEG 300 and 0.01% BC could lead to good nasal delivery systems for OH.
Fumoleau and coworkers115 have carried out randomised, blind, double-dummy, parallel group study of once a day OH suppository (suppository group) with the recommended OH treatment, i.e. 8 mg intravenous (I.V.) OH on day 1 followed by 8 mg tablet (p.o.) twice daily (i.v. + p.o. group) on days 2 and 3 in patients receiving cisplatin (>50 mg/m 2) containing chemotherapy. In the 420 patients included in the intent-to-treat population, 209 received the 16mg suppository and 211 the i.v. + p.o. treatment. The number of emetic episodes and the nausea score were recorded each day. Based on the primary criterion, both treatments provided good anti-emetic control with 87% of all patients having a complete or major response (0-2 emetic episodes) on day 1 in the suppository group and 92% in the i.v. + p.o. group (P= 0.058). The results of this study showed that both treatments are equivalent in the prevention of cisplatin-containing chemotherapy induced emesis for the primary efficacy criteria and that the OH suppository is efficient and well tolerated and is a suitable alternative to the anti-emetic treatment combining the intravenous and oral routes.
27 and Zofran (GlaxoSmithKline, Brazil, as reference formulation) were evaluated following a single 8 mg dose to 23 healthy volunteers of both genders. The dose was administered after an overnight fast according to a two-way crossover design. It was concluded that two OH formulations are bioequivalent in their rate and extent of absorption.
28
2. Scope and objectives of the present work
Oral ingestion is the traditionally preferred route of drug administration, providing a convenient method of effectively achieving both local and systemic effects. In conventional oral drug delivery systems, there is very little control over the release of the drug. The effective concentration at the target site is achieved by intermittent administration of grossly excessive doses, which in most situations often results in constantly changing, unpredictable and often sub- or supra-therapeutic plasma concentrations leading to marked side effects. Also, peroral administration of drugs has disadvantages such as hepatic first-pass metabolism and enzymatic degradation within the gastrointestinal (GI) tract that prohibits oral administration of certain classes of drugs.
With a view to overcome these problems, the current trend in pharmaceutical research is to design and develop new drug delivery systems so as to enhance the therapeutic efficacy of existing drugs. Moreover, the impetus for research in this area can be attributed to the exorbitant cost and large development period involved for ‘new drug development’ when compared to the therapeutic advantages of newer drug delivery technologies. Sustained release technology of drugs has rapidly emerged over the past two decades as a new interdisciplinary science that offers novel approaches to deliver bioactive agents into the systemic circulation. The choice of the drug to be delivered, clinical needs and drug pharmacokinetics are some of the important considerations in the development of such formulations.
29 because of its hydrophilic gel-forming property, non-toxicity and cost effectiveness 40-42
. The swelling rate and erosion of HPMC-based matrix tablets in aqueous media are very crucial in terms of achieving the desired release profiles, and are affected by parameters such as the physicochemical properties of the polymer and the drug, processing conditions, the testing medium used and the formulation composition43–45. The uses of hydrophilic polymers such as hydrogels or pseudo-hydrogels for sustained drug delivery have, therefore, attracted the attention of investigators in recent years. Among the hydrophilic polymers, polysaccharides are preferred due to their nontoxicity and acceptance by the regulating agencies. Polysaccharides, upon contact with water, get hydrated and form gels. When drugs are loaded into these hydrogels, water is absorbed into the matrix, polymer chain relaxation occurs and drug molecules are released through the spaces or channels within the hydrogel network. Polysaccharides like cellulose ethers, xanthan gum, scleroglucan, locust bean gum and gaur gum are some of the natural polysaccharides that have been evaluated in hydrophilic matrix for drug delivery systems117.
Considerable research carried out on locust bean gum105, guar gum118, and tamarind gum119 reveals that polysaccharides can be isolated from their seeds or whole fruits. Delonix regia belongs to the same type containing the pod type fruits. These fruits are dark brown in colour and can be up to 60 cm long and 5 cm wide. The individual seeds, however, are small weighing around 0.4 g on average. These fruits have not been reported so far as a source of polysaccharide for sustained release delivery. Delonix regia was, therefore, chosen in the present investigation for isolation and evaluation of a polysaccharide.
30 rind type fruits. It was, therefore, proposed to investigate the rinds of Artocarpus integrifolia for isolation and evaluation of a polysaccharide.
Considerable research has been carried out on polysaccharides isolated from stem exudates for their pharmaceutical benefits namely, karaya122, acacia123, etc. These polysaccharides have shown good applications in pharmaceutical industries. It was, therefore, proposed to investigate Tamarindus indica and Moringa oleifera exudates for isolation and evaluation of polysaccharides.
In short, in the present investigation, it was proposed to isolate polysaccharides from the plants namely, Artocarpus integrifolia (jack fruit), Tamarindus indica (tamarind), Moringa oleifera (drumstick) and Delonix regia (gulmohar), evaluate them for their sustained release potential and select the best among them for developing SR tablets for certain selected drugs.
The objectives of the present study were, thus, the following;
a. Isolation, purification and characterization of a polysaccharides from Artocarpus integrifolia, Tamarindus india, Moringa oleifera, and Delonix regia and select the best among them for use as a sustained release agent.
b. Toxicity studies and anti-microbial studies on the selected polysaccharide.
c. Preparation of sustained release matrix tablets using the polysaccharide isolated in addition to hydroxypropyl methyl cellulose and xanthan. d. Preparation of sustained release spheroids using the isolated
polysaccharide, xanthan, hydroxypropyl methyl cellulose. e. In vitro dissolution studies.
f. Bioanalytical method development for the selected drug candidates. g. Bioavailability studies in animals and humans.
2.1 Selected drugs
31 induced by cytotoxic chemotherapy and radiotherapy is due to antagonism of 5HT3 receptors on neurons located both in the peripheral
and central nervous system124. It has a half-life of ~ 5.7 h with oral dosage regimen of 8-32 mg 4 times a day125.
C H3
N
N N
C H3
O
Figure 3. Structure of OH
OH is passively and completely absorbed following oral administration from the gastrointestinal tract and undergoes first pass metabolism. The most common side effect is headache and the other side effects include seizers, extrapyramidal effects such as dystonic reactions and dyskinesia.
32 upset stomach, dizziness, a stuffy nose and substantial weight gain or weight loss are the less common side effects127-129.
S N
N
N
O
OH
Figure 4. Structure of QF
2.2 Proposed plan of work
The project was proposed to be carried out in the following stages;
Step I: Isolation, purification and characterization of polysaccharides 1.Collection and authentication.
2.Purification of the gum 3.Characterization of the gum
• Identification • Bulk density
• Solubility • True density
• Color and clarity • Porosity of powder
• pH • Density and Specific gravity
• Swelling index • Powder fineness • Loss on drying • Ash values • Hygroscopicity • Microbial count • Angle of repose • Dissolution studies
Step II: Toxicity studies
1. Acute Oral Toxicity Study (Up and Down Method) of the selected (Delonix regia) polysaccharide in Wistar Rats
33 Step III: Preformulation studies
1. Determination of physical properties of the drugs such as physical nature (amorphous or crystalline) solubility, melting point, etc.
2. Development of calibration curve for the selected drugs 3. Compatibility studies by
• IR spectral matching approach
• Differential Scanning Calorimetry (DSC studies)
Step IV: Development of oral sustained drug delivery systems 1. Development of matrix tablets by wet granulation
Formulation and characterization of granules for, • Angle of repose
• Loose bulk density • Tapped bulk density • Compressibility index • Drug content
Compression of the formulated granules into tablets and evaluation of the tablets as per the pharmacopeial specifications for,
• Average weight and weight variation • Thickness
• Diameter
• Drug content and content uniformity • Hardness
• In vitro drug release behavior and comparison of the release profile with the marketed conventional dosage forms.
2. Development of spheroids by extrusion spheronization technique Formulation of spheroids by wet granulation technique and evaluate them for
34 • In vitro drug release behavior and comparison of the release profile
with marketed formulation.
Step V: Stability studies as per the ICH guidelines
Stability studies of the selected formulations at the following temperature and humidity conditions as prescribed by the International Conference on harmonization (ICH).
• 25 °C with 60 % RH • 40 ° C with 75 % RH
Step VI: Development and validation of HPLC method Optimization of the chromatographic conditions likes,
• selection of wavelength,
• selection of initial separation conditions,
• nature of the stationary phase,
• nature of the mobile phase (pH, peak modifier, solvent strength, ratio and flow rate),
• sensitivity and
• Selection of internal standard.
Validation of the developed methods using the various validation parameters such as,
• Accuracy,
• Precision,
• Linearity and Range,
• Limit of detection (LOD) / Limit of quantitation (LOQ),
• Selectivity / specificity,
• Robustness / ruggedness,
35 Step VII: Bioavailability studies
Carry out bioavailability studies in a cross over design in New Zealand albino rabbits / healthy human volunteers between the developed formulation and the conventional dosage form.
The following pharmacokinetic parameters were proposed to be evaluated and compared between the formulations developed and the conventional IR dosage forms.
• Cmax Maximum plasma concentration
• tmax Time of maximum plasma concentration
• AUC0-t Area under plasma concentrations time curve 0 to 24 h • AUC0- ∞ Area under plasma concentrations time curve 0 to ∞ h
• t1/2 Elimination half-life • kel Elimination rate constant
2.3 Plant profiles
(i) Delonix regia (Gulmohar)
Delonix regia is a flowering plant from the Fabaceae family, noted for its fern-like leaves and flamboyant display of flowers. Often grown as an ornamental tree and given the name Royal Poinciana or Flamboyant, it is also known as Krishnachura, Gulmohar, Peacock Flower, Flame of the Forest and Malinche101 In addition to its ornamental value, it is also a useful shade tree in tropical conditions, because it usually grows to a modest height (typically around 5 m, though it can reach as high as 12 m) but spreads widely, and its dense foliage provides full shade. Seed pods are dark brown and can be up to 60 cm long and 5 cm wide; the individual seeds, however, are small, weighing around 0.4 g on average. In India, it is referred to as the Gulmohar. In West Bengal (India) and Bangladesh it is called Krishnachura.
36 Pods are large, flat, 40 to 70 cms by 2.5 to 4 cms in size, compressed, hard, brown or black when ripe. 20 to 30 seeds in distinct cavities, grayish white, with a dark ridge, hard bony testa, oblong, 1.5 to 2 cms in length.
(ii) Artocarpus heterophyllus (Jackfruit Waste)
Jack fruit tree (Artocarpus heterophyllus Lam.) belongs to Moraceae family. It is a medium-sized tropical fruit tree reaching 15-20 m in height. The evergreen leaves are oblong, oval or elliptic in shape, 10-15cm in length, alternate, glossy and dark green in colour. The juvenile leaves are lobed. The tree is monoecious, producing male and female flowers. The male flowers are produced amongst the leaves above the female flowers, and when mature, become covered in pollen that falls rapidly after flowering. The female flowers are borne on short twigs that develop from the trunk, branches and sometimes from below the soil level at the base of older trees.
Jackfruit is the largest tree-borne fruit in the world, reaching up to 50 kg in weight and 60-90 cm in length. A mature tree produces up to 700 fruits per year, each weighing 0.5 to 50 kg. The rind of the compound fruit is greenish yellow when fully ripe. Inside, the fruit is made up of large, yellow bulbs enclosing an oval light-brown seeds. There are 100-500 seeds in a single fruit. When fully ripe, the opened jackfruit smells of pineapple and banana. All parts of the tree produce sticky, white latex, but gum-free genotypes have been identified in India. Edible bulbs of ripe jackfruit are consumed for their fine taste and pleasant aroma. The edible portion is about 30% by weight130.
About 50% of fruit is composed of rind and unfertilized floral parts, which are also rich in jackfruit flavor, are usually discarded as waste, because they are fibrous1. Cattle feed showed the non-edible portion (perianth meal, rind and core meal) of the fruit was 59·2% with a total dry meal recovery of 11·6%131.
(iii) Tamarindus indica (Tamarind)
37 pedicels 8. 10 mm, calyx circa 1.2 cm long, tube turbinate, teeth lanceolate, the lowest two connate. 1 - 3 petals with 1 - 1.5 cm length, pale yellow with red veins. Pods 7.5 to 20 cm long, 2-2.5 cm wide, slightly compressed, indehiscent. Seeds are dark brown or black.
The tamarind tree is a host for the lac insect, Kerria lacca that deposits a resin on the twigs. This product could be harvested and sold as stick-lac for the production of varnishes and lacquers. However, this is not considered an important product and tamarind growers often cut the resinous twigs and discard them.
The fruits are pods 5-10(-16) cm long x 2 cm broad, oblong, curved or straight, with rounded ends, somewhat compressed and indehiscent although brittle. The pod has an outer epicarp which is light grey or brown and scaly. Within is the firm but soft pulp which is thick and blackish brown. The pulp is traversed by formed seed cavities, which contain the seeds. The outer surface of the pulp has three tough branched fibres from the base to the apex. Each pod contains 1-12 seeds which are flattened, glossy, and orbicular to rhomboid, each 3-10 x 1.3 cm and the centre of each flat side of the seed marked with a large central depression. Seeds are hard, red to purple brown, non arillate and exalbuminous. Seed chambers are lined with a parchment like membrane. Cotyledons are thick. Pods ripen about 10 months after flowering and can remain on the tree until the next flowering period, unless harvested 132, 133.
Moringa oleifera (Drumstick)
Moringa oleifera Lam. belonging to the single- genus family Moringaceae is a small fast-growing ornamental tree widespread over the tropical regions of Africa and Asia134. Moringa has its origin in Arabia and India but today it is common all over the tropics, from South Asia to West Africa. Moringa is found in parts of East and South Africa, in many Pacific Islands in South and Central America.
38 Moringa does best where the temperature ranges from 25 to 40 degrees C (77 to 104 degrees F) and annual rainfall is at least 500 mm. Moringa grows well from sea level to 1,000 m (3,000 ft.) elevation. Historically, there is evidence that cultivation of Moringa in India dates back thousands of years, and the traditional Ayurvedic medicine used this tree to heal or prevent hundreds of diseases.
The fruits are pendulous, linear, three-sided pods with nine longitudinal ridges, usually 20 to 50 cm long, but occasionally up to 1 m or longer, and 2.0 to 2.5 cm broad. The pods, each usually containing up to 26 seeds, are dark green during their development,and take approximately 3 months to mature after flowering90. They turn brown on maturity, and split open longitudinally along the three angles, releasing the dark brown, trigonous seeds. Seeds measure about 1 cm in diameter, with three whitish papery wings on the angles.
F-1
Table 1. Seed biology of Delonix regia
No. of seeds / kg Purity
(%)
Moisture
(%)
Germination
(%)
Plant
(%)
No of seedlings / kg.
of seed
2910 to 3245 100 6.4 24 to 68 19 to 30 420 to 975
F-2
Figure 5. Delonix regia. A. Whole tree. B. Pod with flower. C. Seeds in the pod.
D. Transfer section of the seed. D
F-3
Figure 6. Artocarpus heterophyllus. A. Transverse section of the whole fruit. B & C. Edible portion in the non edible portion.
A
C
B
39
3. Experimental
3.1. Materials
Ondansetron hydrochloride (OH) was procured from Microchem Services, Bangalore, India. Quetiapine fumarate (QF) was procured from Beijing Lunarsun Pharmaceutical Co., Ltd, China. Methocel –K-100 CR (HPMC–K-100 CR, Dow chemicals, USA) and starch 1500 were gift samples from Colorcon India. Xanthan gum was a gift sample from Lupid Colloids, Mumbai. Gulmohar seeds and moringa gum were collected in Kumbakonam, Middle eastern part of Tamilnadu, India. Tamarind gum was purchased from Abirami Enterprises, Chennai, Tamilnadu, India. The jack fruit wastage was collected from the local market, Ooty, Tamilnadu, India. Avicel PH 101 [Microcrystalline cellulose (MCC) from FMC polymers, Ireland] was procured from Signet chemicals, Mumbai. PVP-K- 30 (Poly vinyl pyyrolidine from ISP) was a gift sample from Anshul agencies, Mumbai. Lactose I.P and Aerosil (colloidal silicon di oxide) were purchased from Lactose India Pvt. Ltd, Thane and Degussa, Mumbai, respectively. All the reagents and chemicals used for analytical development were of HPLC grade.
The polysaccharides were isolated from different plant source, namely, Artocarpus integrifolia, Tamarindus india, Moringa oleifera and Delonix regia.
3.2. Isolation of the polysaccharide from Moringa oleifera (Moringa gum)
40 process of dissolution in water and precipitation with alcohol was repeated six times until an almost white precipitate was obtained90. The dried polysaccharide was milled and sifted with an 80 mesh sieve to obtain a flour (MG flour).
3.3. Isolation of the polysaccharide from Tamarindus indica (Tamarind gum) The gum was thoroughly cleaned to remove all the adhered foreign materials and powdered. It was then washed several times with alcohol. 100g of the gum was soaked in 1L of warm water for overnight. It was then milled by stirring vigorously, boiled for 2-4h, cooled to room temperature and squeezed using several folds of muslin cloth to separate the marc from the filtrate. The marc was discarded. An equal volume of absolute alcohol was added to precipitate to the filtrate the polysaccharide. It was then separated by filtration and dried in an oven at 60oC. The brown color gum was collected, washed several times with ethanol and dried. The process of dissolution in water and precipitation with alcohol was repeated until an almost white precipitate was obtained91. The dried polysaccharide was milled and sifted with an 80 mesh sieve to obtain a flour (TG flour).
