Accepted Manuscript

Title: Physicochemical and release properties of

carboxymethylated starches of Dioscorea from Jharkhand

Author: Deepika Vashisht Anima Pandey K. Jayaram Kumar

PII: S0141-8130(14)00798-3
DOI: http://dx.doi.org/doi:10.1016/j.ijbiomac.2014.11.039
Reference: BIOMAC 4756

To appear in: International Journal of Biological Macromolecules

Received date: 5-10-2014

Revised date: 12-11-2014
Accepted date: 21-11-2014

Please cite this article as: D. Vashisht, A. Pandey, K.J. Kumar, Physicochemical
and release properties of carboxymethylated starches of Dioscorea from
Jharkhand, International Journal of Biological Macromolecules (2014),
http://dx.doi.org/10.1016/j.ijbiomac.2014.11.039

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1 Physicochemical and release properties of carboxymethylated starches of Dioscorea from

2 Jharkhand

3 Deepika Vashisht, Anima Pandey, K Jayaram Kumar*

4 Department of Pharmaceutical Sciences & Technology, Birla Institute of Technology, Mesra,

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5 Ranchi-835215, Jharkhand, India.

6 * Corresponding author. Tel.: +91 06512276247; fax: +91 06512275290.

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7 E-mail addresses: jayarampharm@gmail.com

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8 (K. Jayaram Kumar).

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 Abstract

9 Increasing demand and considerable attention to the non-conventional sources of starches leads

10 to explore new sources. Starches of Dioscorea (Da1 and Da2) from Jharkhand, North Eastern

11 region of India have been studied for its physicochemical properties. An attempt has been made

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12 to study the carboxymethylated derivatives of starches from two varieties of Dioscorea of this

13 region. Different concentration of monochloroacetate was used to study the effect of degree of

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14 substitution (DS) on the physicochemical properties of starches. A considerable effect of DS was

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15 noticed on the ash content, amylose content, water-holding capacity, swelling and solubility

16 power of carboxymethylated derivatives. Morphological studies showed the increase in the

17

18 an
deformation of structure of the starch granule with an increase in the degree of substitution. FTIR

confirmed the carboxymethylation reaction. TGA data of carboxymethylated starches revealed

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19 the stability to the temperature. Micromeritics of starch powder and granules showed the value
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20 which makes these starches to be utilized as an excipient. With the increase in DS, the % release
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21 of drug was found to be decreased. This further makes the carboxymethyl derivatives of

22 Dioscorea a good source to be used as an excipient for sustained release formulations.

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23 Keywords: Dioscorea, Carboxymethylation, Swelling power, Sustained release

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Page 2 of 26
 1. Introduction

24 Starch is one of the suitable, compatible and cheapest biopolymer as compared to other

25 biomaterial resources, used as matrix forming material in pharmaceutical, food, textile and paper

26 industries [1]. Starch in its innate form has many disadvantages related to hydrophilicity,

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27 mechanical properties and stability, which make it undesirable for aforesaid industries. One of

28 the possible ways to overcome these limitations and to extend the asset of starch are its

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29 modifications through physical and chemical means [2-4]. Physical modifications comprise heat

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30 moisture treatments, retrogradation, annealing, etc. [3, 5, 6] while chemical modifications can be

31 achieved through carboxymethylation, acetylation, succinylation, esterification, etherification,

32

33
etc. [7, 8].

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Chemical modification involves the derivatization or decomposition reactions which incorporate
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34 functional group to the starch [8]. Chemical modification improves the working properties of the
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35 starch and customized it to specific applications. Carboxymethylation is one the chemical

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36 modification gaining attention due to its various applications in industries. Carboxymethyl starch

37 (CMS) is used as ion sorbent [9], sizing agent [10, 11], thickener [12], finishing agent [13] etc.
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38 Due to increased solubility, CMS increased the suspending and cleaning capability of detergent
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39 [14].
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40 CMS also have acquisition in pharmaceutical formulation for sustained release drug [15] and it

41 has been reported that CMS increase the drug loading capacity upto 60% [16]. Different studies

42 on carboxymethylation of starches from distinct sources have been reported including maize,

43 potato, cassava, arrow root, mung bean, etc. But to the best of our knowledge

44 carboxymethylation of Dioscorea starch from North Eastern region of India has not been studied

45 yet. The aim of the present study is to synthesize and characterize the carboxymethylated starch

Page 3 of 26
46 of two varieties of Dioscorea (Da1 and Da2) and to study the effect of degree of substitution on

47 the physicochemical and release properties of carboxymethylated starches.

48 2. Materials and Methods

49 2.1 Materials

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50 Tubers of Dioscorea were obtained from local market of Jharkhand and these were termed as

51 Da1, Da2. Reagents and chemicals used throughout the experiments were of analytical range.

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52 2.2 Isolation of starch

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53 Starches from both the tubers were isolated by using the methodology as described by Flores-

54 Gorosquera et al. [17] and Deepika et al. [18]. For isolation of starch, tubers of Dioscorea were

55

56 an
thoroughly washed, peeled and cut into pieces. Pieces were then soaked into citric acid (0.3%,

w/w) to remove the mucilage from starch and milled in a blender using distilled water. Paste was
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57 thoroughly washed with water until washing water become clear. Suspension was allowed to
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58 settle and after settling water was removed. Crude starch was then dried in air and stored in
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59 polythene bags for further study.

60 2.3 Carboxymethylation of isolated starches

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61 Carboxymethylation of starches from both varieties of Dioscorea (Da1 and Da2) was achieved by
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62 the method as reported by the Afolabi (2011) [19] and modified method of Kulkarni et al. (2013)
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63 [20]. Weighed quantity of NaOH (3.2 g) was mixed to 20 mL of water in a round-bottom flask,

64 and mixture was stirred continuously until the alkali dissolved completely. About 125 mL of

65 isopropyl alcohol was added to the alkali solution and temperature was increased to 40 °C.

66 Starch (8 g) was added to the solution and stirred with the nitrogen gas flushed throughout the

67 reaction. After 1 h different quantities monochloroacetate (MCA; 5 g, 10 g and 15 g) was added

68 to the mixture, and reaction was continued for another 4 h. Resultant mixture was filtered, taken

Page 4 of 26
69 in methanol and neutralized with acetic acid. The suspended was washed with aqueous methanol

70 (80%) several times to remove unreacted reagent. The slurry then dispersed in acetone, stirred

71 for 20 min and dried in hot air oven at 40 °C to obtain carboxymethylated derivative termed as

72 Da1CS1, Da1CS2, Da1CS3 for Da1 variety and Da2CS1, Da2CS2, Da2CS3 for Da2 variety with

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73 respect to the three different concentration of monochloroacetate.

74 2.4 Degree of substitution (DS) and Reaction Efficiency

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75 Back titration was considered to be the most appropriate method to determine the degree of

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76 substitution (DS) [21]. About 0.50 g of CMS was taken in 20 mL of NaOH (0.2 M) followed by

77 addition of 50 mL of distilled water. The solution was then transferred to a 100 mL volumetric

78

79 an
flask, and volume was made up to 100 mL with distilled water. From the resultant solution, 25

mL was taken in an Erlenmeyer flask and 50 – 100 mL of distilled water was added to dilute it.
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80 Excess of NaOH was back titrated with standardized 0.05 M HCl to a phenolphthalein endpoint.
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81 Native starches were also treated in the same way to get the value for blank. DS was calculated
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82 using the following formula [22]:

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83
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84 (1)
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85

86 (2)

87 here, 162 g mol−1 = molecular weight of anhydroglucose unit; 58 g mol−1 is the net increase in

88 the mass of an anhydroglucose unit (AGU) for each carboxymethyl group substituted

89 Reaction efficiency (RE) [23, 24], was computed by the equation

90 (3)

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 91 (4)

92 DSt is the theoretical degree of substitution (DS); nAGU the number of moles of AGU; and

93 nNaOH, number of moles of sodium hydroxide.

94

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2.4 Analysis of carboxymethylated starches

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95 2.4.1 Moisture, ash content and elemental analysis

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96 Moisture and ash content of the starch samples were investigated by AOAC methods [25].

97 Briefly, moisture content was determined by heating the samples at 105 °C for three hours in a

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98 tared petridish. Ash content was investigated by igniting the samples at 500 °C for 12 h.

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99 Elements viz carbon, hydrogen, nitrogen and sulphur were deteced by an Elemental Analyzer

100 (Make- M/sElementar, Germany; Model-Vario EL III).

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101 2.4.2 Amylose Content

102 Amylose content was analyzed and estimated by using the method adopted by Singh et. al [26].
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103 Each starch sample was weighed (100 mg) and taken in volumetric flask (100 mL). These
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104 samples were heated with 1mL of 95% ethanol and 9 mL of 1 N sodium hydroxide for 10 min.
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105 Prepared material were then cool down and volume was made upto 100 mL with distilled water.
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106 A part of all the samples (5 mL) was taken in 100 mL volumetric flask, to this 1 mL of 1 N

107 acetic acid and 2 mL of iodine solution was added, and volume was made upto 100 mL using
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108 distilled water. After 20 min. absorbance at 620 nm was measured spectrophotometrically:

109 % Amylose = 3.06 × absorbance × 20 (5)

110 2.4.3 Water holding capacity

111 Water holding of all the modified starches were analyzed by using the method of Yamazaki [27]

112 and modified method of Medcalf and Giles [28]. A suspension was prepared using each starch

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113 (1g) in 15 mL of water and stirred for 1 h. Resulting suspension was then centrifuged at 3000

114 rpm for 10 min. Free water was removed and weight of wet starch was taken.

115 (6)

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116 Here, WtS = weight of wet starch (g), W = initial weight of starch (g)

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117 2.5 Micromeritics properties

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118 Flow properties of the modified starches were assessed in terms of bulk density, tapped density,

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119 Angle of repose, Hausner’s ratio, Carr’s index by using standard methods as mentioned in USP

120 (USP XXXI) [29] and described by Carr [30].

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121 2.6 Swelling power and solubility power

122 A suspension of each starch was heated in a water bath at different temperatures of 30 °C to 90
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123 °C for 30 min with continues stirring to counter the lump formation. Suspension was then

124 centrifuged at 3000 rpm for 15 min. Supernatant was removed cautiously and deposited swollen
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125 starch was weighed. Aliquot of each sample was taken in pre-weighed petridish and these were
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126 dried to constant weight at 105°C. The swelling power (Swell. P, g/g on the dry weight basis)
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127 and solubility power (Sol. P, %) [31] were computed as:

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128
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129 (7)

130 (8)

131 2.8 Morphological Studies

132 Granule morphology of carboxymethyl derivatives was examined under Scanning electron

133 microscope (JEOL – Japan, JSM 6390 LV) placing samples mounted on a metal stub using

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134 double sided adhesive tape. Samples were coated with platinum to make samples enough

135 conductive and images were obtained under an accelerating voltage of 10 kV.

136 2.9 Fourier Transform Infrared Spectroscopy

137 Structural analyses of all the carboxymethylated samples were performed on the FTIR (FTIR-

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138 8400S, Shimadzu, Japan) spectra by recording the transmittance in the range of 4000–400 cm−1.

139 2.10 Thermogravimetry Analysis

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140 Thermogravimetric analysis of derivitized samples was accomplished on DTG-60 (Shimadzu,

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141 Japan). The measurements were carried out in nitrogen atmosphere with a purge rate of 50

142 mL/min by heating the samples from room temperature to 500 °C at a heating rate of 10 °C.

143

144 an
Samples were hold in a platinum crucible and weight loss in percentage was recorded from the

onset temperature measured for each carboxymethyl starch.

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145 2.11 Preparation and evaluation of granules
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146 Wet granulation method of tablet formulation was employed for granule preparation using
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147 paracetamol (PCM) as model drug. Different carboxymethylated starches as binder were

148 incorporated to study the effect of degree of substitution on release profile of drug. PCM (46.8%
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149 w/w), starch (4.7% w/w), lactose (28.8% w/w) and sodium CMC (7.2% w/w) was blended with
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150 water, used as granulating liquid, to form a damp coherent mass, and it was passed through sieve
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151 20# to form granules. Talc and magnesium stearate were used as a lubricant. The granules were

152 evaluated for bulk density and tapped density, angle of repose, Hausner’s ratio and

153 compressibility index [29, 30].

154 2.12 Manufacture of tablets and evaluation of tablets

155 Paracetamol tablets were manufactured from prepared granules by compressing them in a 16

156 station tablet punching machine (Cadmach, Ahmadabad, India). Prepared tablets were selected at

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157 random from each batch and evaluated for elastic recovery, weight variation, hardness, % drug

158 content, and friability [29].

159 2.13 In-vitro release profile of tablets prepared using carboxymethylated starches

160 Release profile of each tablet was performed in a USP Type-II dissolution test apparatus

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161 (Electrolab, Mumbai, India) according to the method mentioned in USP (USP XXXI). Studies

162 were carried out in 900 mL of different dissolution media of pH 1.2 depicts gastric fluid for first

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163 2 h, pH 6.8 represents simulated intestinal fluid for next 2 h and pH 7.4 for next 4 h. At the

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164 predetermined time intervals five mL of samples was withdrawn and immediately replaced with

165 fresh medium to maintain constant volume. Amount of paracetamol was determined by

166

167 3. Results and Discussion an

measuring the absorbance at 243 nm spectrophotometrically.
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168 3.1 Proximate analysis of Derivative starches
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169 Degree of substitution (Table 1) in all the derivatives was found to be increased with an increase
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170 in the concentration of monochloroacetate. DS of the entire carboxymethyl derivative was found

171 in the range of 0.43-1.87. Similar increment in the RE was observed in all the samples. This
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172 increment in DS may be ascribed to increase in the interaction between starch granules and the
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173 etherifying reagent when concentration of MCA increased. Nevertheless, increment in the
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174 concentration of MCA can possibly cause side reaction, which may decrease DS and RE [32-

175 34]. Moisture content for the carboxymethyl derivatives was ranged from 8.09% - 12.32%, and it

176 is in the range 10% and 17% as mentioned in the literature [35, 36]. The highly substituted

177 samples were found to have more ash value and water-holding capacity as compared to the

178 native samples as reported earlier [18]. Amylose content was observed to be a decrease with the

179 increase in substitution in all the samples, and it was ranging from (21.16 % -22.36 %).

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180 Elemental analysis (Table 1) showed the less amounts of nitrogen and sulphur, which are an

181 indication of purity of starches.

182 [Insert Table 1]

183 3.2 Micromeritics Properties

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184 Micromeritics of the derivatives of Dioscorea starches (Da1 and Da2) has been given in Table 2.

185 Carboxymethylation of the native Dioscorea starches found to have considerable effect on the

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186 flow properties of the starches as compared to be previously reported native starches [18]. This

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187 proves that carboxymethylation could improve the flowability of starches during compression.

188 Powders with Hausner’s ratio less than 1.25 have good flow properties [37] and granules with

189

190 an
angle of repose less than 30º show good flow [38]. Higher content of water have been shown to

affect the flow and mechanical properties of starches as revealed by the thermogravimetric
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191 analysis. All the samples were having the flow parameters within the range and Hausner’s ratio
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192 and angle of repose indicated the good flow properties of samples (Table 2).
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193 [Insert Table 2]

194 3.3 Swelling and Solubility Power

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195 The results showed improved swelling power (Table 3) following carboxymethylation. The
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196 swelling power of the carboxymethylated derivatives of Dioscorea starches was varied from
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197 6.20% – 23.77% with the increase in the temperature. This increase might be due to the

198 weakening of forces between the granules upon modification, which allow the penetration of

199 water and subsequently increase in swelling power [22]. This can also be well supported by the

200 thermal studies of the carboxymethylated starches. This can also be attributed to inclusion of the

201 carboxyl group to the starch which resulted in loss of crystallinity and increase the amorphous

202 region [20]. Solubility power (Table 3) of the derivitized Dioscorea starches was found to

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Page 10 of 26
203 decrease as compared to the previously reported their native forms [18] but found to increase

204 with an increase in temperature. At the higher temperatures the mobility of starches increases

205 which increase the dispersion of the starch granules in the solution and hence improve the

206 solubility. Reduction in solubility might be due to the addition of the bulky carboxyl group to the

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207 starches which subsequently decreases the mobility of starch and hence results into the decrease

208 in solubility [39].

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209 [Insert Table 3]

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210 3.4 Morphological Studies

211 Granules of carboxymethylated starches were observed under scanning electron micrograph and

212

213 an
micrographs of the starches were sown in Fig. 1. From the scanning micrographs, it is clear that

carboxymethylation altered the granule morphology. The granules were found more deformed as
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214 the DS increased. This deformation can be correlated to the availability of strong alkali
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215 environment during carboxymethylation. Granules of native starches as reported earlier [18]
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216 were well defined ellipsoidal shape but modification results into alteration of crystallinity of

217 starch. This alteration in the morphology allows the more interaction of starches with the
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218 etherifying agent for reaction to occur and distorted the structure.
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219 [Insert Fig. 1]

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220 3.5 FTIR spectral studies

221 The introduction of the carboxyl group in the native starches can be proved by FTIR spectral

222 studies. Fig. 2A and 2B displaying the FTIR analysis of carboxymethylated Dioscorea starches.

223 In all the modified samples the peaks near 970 cm-1 and 1200 cm-1 are preserved, which are

224 typical peaks of starch [40] and proves the carbohydrate nature of the samples. The band near

225 2900 cm−1 is assigned to -CH2 symmetrical stretching vibrations. Incorporation of the carboxyl

11

Page 11 of 26
226 group can be indicated by new peaks observed near at 1600 cm-1, which corresponded to –COO˗

227 asymmetric stretching vibrations [41]. Peaks at 2900 cm-1 and 2400 cm-1 may be due to the CH2

228 asymmetric stretching [19]. A broad band at 3200 – 3400 cm-1 was observed in native starches

229 and it was found to decrease, which further is an indication of carboxymethylation reaction [19].

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230 This broad band attributed to the number of hydrogen bonded-hydroxyl group, which were found

231 to reduce after carboxymethylation. This reduction may be due to the incorporation of carboxyl

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232 group to the starch [19].

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233 [Insert Fig. 2A and 2B]

234 3.6 TGA studies of carboxymethylated derivatives

235

236 an
Thermal stability of the carboxymethylated Dioscorea starches was observed by TGA (Fig. 3A

& 3B). Thermo-physical criteria specified about the thermal stability of polymers. Such
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237 information is required for acceptable use of polymers in various industrial applications [42].
d

238 During heating weight loss was occurred in two steps decomposition in both native and
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239 carboxymethylated starches. The native starches and carboxymethylated starches the first

240 decomposition at 250 °C. The weight loss occurred in first step was ranged from 8.99% to
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241 18.94%. This could explain the improved swelling kinetics and solubility of the
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242 carboxymethylated derivatives of starches up on increase in temperature. The first event of

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243 weight loss is due to the degradation of water molecules [43]. There is 67-72% loss was occurred

244 for native starches in second step of decomposition, and in case of carboxymethylated starches

245 67 – 79% loss occurred, which is near to the native starches. At 500 °C percent weight loss for

246 all the carboxymethylated derivatives was less than the native starches i.e. it was found in the

247 range 76-89% and this data represents that carboxymethylation mended the thermal stability of

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Page 12 of 26
248 the samples. This is due to the fact of replacement of the hydroxyl group with carboxymethyl

249 during the carboxymethylation.

250 [Insert Fig. 3Aand 3B]

251 3.7 Micromeritics properties of granules and physical properties of tablets

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252 Granules prepared by using derivitized starches showed the values (Table 4) within the range as

253 mentioned in the USP (USP XXXI) [29]. These values of densities in the table 4 indicate the

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254 more close packing and better die filling during the compression of granules which is further

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255 confirmed by Carr’s index and Hausner’s ratio. The granule showed the Carr’s index between

256 12.50% - 16.94% and Hausner’s ratio 1.14-1.20. The hardness of the modified starches was

257

258 an
found as the value above the 4 kg/cm2. The hardness should be between 4-6 kg/cm2. The

friability of all the formulation was less than 1%, which meets the values specified in USP (USP
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259 XXXI) [29]. With the increase in DS hardness and friability of tablets was found to increase. The
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260 results mentioned that carboxymethylation led to improvement in the physical properties of
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261 starches, which may further affect the release properties. The elastic recovery of the tablets was

262 also within the limits which are the indication of better compressibility. Drug content was found
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263 in the varied from 98.25% - 100.21%, which meets the specifications mention in USP (USP
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264 XXXI) [29].

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265 [Insert Table 4]

266 3.8 In-vitro release profile

267 All the modified starches were studied for their release profile (Fig. 4) in the physiological

268 environment. It was found that with an increase in the DS, the % release decreases in both the

269 varieties Da1 and Da2. There is only 75.95 % and 82.19 % release in case of Da1CS3 and Da2CS3

270 respectively in 8 hrs. The decrease in release rate may be due to the less swelling of starch below

13

Page 13 of 26
271 50 °C [44]. The drug release rate can be affected by water permeation, polymer swelling and

272 erosion of the matrix [45]. Further from the data we can say that carboxymethylated tablets

273 remain intact throughout the dissolution process and showed less release rate. These studies

274 prove the effect of DS on the release profile of drug and also justified that the carboxymethylated

t
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275 derivatives can be a good prospect in sustained release formulations.

276 [Insert Fig. 4]

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277 4. Conclusion

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278 The present study revealed the utility of carboxymethylated starches of Dioscorea as excipient in

279 pharmaceutical industries. DS was found to have an impact on the various properties of

280

281 an
derivitized starches. The results suggested the better flow properties of the derivatives which

were in the acceptable range. The swelling power was found to be substantially increased with
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282 increase in the degree of substitution corresponding to the temperature. The derivatives of
d

283 Dioscorea starches produced the granules with good flow properties and tablets with
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284 considerable physical properties. The carboxymethylation of starches imparted the hydrophilicity

285 to the starch which significantly prevents the release of drug in dissolution media. The present
p

286 study proves the use of the carboxymethyl derivative of Dioscorea starches for sustained release
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287 drug formulations.

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288 Acknowledgements

289 The authors are grateful to Council of Scientific and Industrial Research (CSIR), New Delhi to

290 provide a grant to carry out this study. We thank to the Central Instrumentation Facility, BIT

291 Mesra for providing instrumentation facilities.

292 References

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327 [27] W.T. Yamazaki, Cereal Chem. 30 (1953) 242–246.
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p

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334 [34] O.S., Lawal, M.D. Lechner, H. Brigitte, W.M. Kulicke, Starch/ Stärke 59 (2007) 224–233.

335 [35] G. Rowley, Starch, in: R.C. Rowe, P.J. Sheskey, P.J. Weller (Eds.), Handbook of

336 Pharmaceutical Excipients, Pharmaceutical Press, London, 2003, pp. 603–608.

337 [36] H. Leach, Gelatinization of starch, in: R.L. Whistler, E.F. Paschall (Eds.), Starch:

338 Chemistry and Technology, Academic Press, New York, 1965, pp. 289–308.

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339 [37] D. Panda, N.S.K. Choudhury, M. Yedukondalu, S. Si, R. Gupta, Indian J. Pharmac. Sci.

340 70 (2008) 614- 618.

341 [38] K. Reddy, S. Mutalik, S. Reddy, AAPS Pharm. Sci. Tech. 4 (2003) 480-488.

342 [39] O.S. Kittipongpatana, N. Kittipongpatana, LWT-Food Sci. Technol. 44 (2011) 1766–

t
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343 1773.

344 [40] M.I. Khalil, A. Hashem, A. Hebeish, Starch/Stärke 42 (1990) 60–63.

cr
345 [41] H. Qiu, L. He, Polym. Adv. Technol. 10 (1999) 46 8–472.

us
346 [42] K. P. Menard In: Brostow W, (Eds.) Performance of plastics. Hanser: Munich–Cincinnati;

347 2000.

348

349 Polym. 88 (2012) 299–307. an

[43] J. Castano, R. Bouza, S. Rodríguez-Llamazares, C. Carrasco, R.V.B. Vinicius, Carbohydr.
M
350 [44] C. Freire, F. Podczeck, F. Veiga, J. Sousa, Eur. J. Pharm. Biopharm. 72 (2009) 587–594.
d

351 [45] B. Bharaniraja, Jayaram K. Kumar, C.M. Prasad, A.K. Sen, Int. J. Biol. Macromol. 49
te

352 (2011) 305–310.

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 t
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353 Table 1 Proximate analysis of carboxymethyl derivatives of Dioscorea starches (Da1 and Da2)

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Samples DS % RE Moisture (%) Ash Amylose (%) Water Holding Elemental Analysis

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Capacity (%) C H N S

Da1CS1 0.43 32.9 8.09 ± 0.01 0.78 ± 0.02 21.20 ± 0.02 438 ± 0.02 35.31 6.848 0.279 0.361

an
Da1CS2 1.01 50.6 11.21 ± 0.02 0.81 ± 0.05 21.19 ± 0.02 510 ± 0.02 37.91 7.249 0.450 0.188

Da1CS3 1.87 79.6 11.26 ± 0.01 0.80 ± 0.02 21.16 ± 0.11 517 ± 0.03 38.13 7.472 0.258 0.140

M
Da2CS1 0.53 37.3 11.89 ± 0.01 0.58 ± 0.15 22.36 ± 0.02 579 ± 0.02 35.94 6.986 0.010 0.074

0.89 54.7 12.30 ± 0.05 0.58 ± 0.02 22.22 ± 0.12 582 ± 0.05 36.74 7.428 0.000 0.033

ed
Da2CS2

Da2CS3 1.28 59.9 12.32 ± 0.01 0.60 ± 0.02 22.20 ± 0.02 596 ± 0.02 36.94 7.662 0.014 0.043
pt
All the data were represented as average of triplicate analysis ± standard deviation.
354

DS = Degree of substitution; % RE = Reaction efficiency

355
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 t
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356 Table 2 Powder characteristics of carboxymethylated derivatives of Dioscorea starches (Da1 and Da2)

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Samples Bulk Density Tapped Density Hausner’s Carr’s Index Angle of Repose Porosity (%)

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(g/cm3) (g/cm3) Ratio (%) (°)

Da1CS1 0.36 ± 0.01 0.39 ± 0.12 1.11 ± 0.01 7.69 ± 0.06 17.24 ± 0.15 77 ± 0.01

an
Da1CS2 0.42 ± 0.02 0.47 ± 0.15 1.13 ± 0.21 10.63 ± 0.01 18.54 ± 0.01 78 ± 0.08

Da1CS3 0.38 ± 0.05 0.44 ± 0.01 1.16 ± 0.04 13.63 ± 0.11 19.36 ± 0.17 76 ± 0.01

M
Da2CS1 0.51± 0.01 0.61 ± 0.05 1.21 ± 0.01 16.39 ± 0.20 20.85 ± 0.01 74 ± 0.18

0.48 ± 0.02 0.56 ± 0.07 1.17 ± 0.15 14.28 ± 0.01 22.63 ± 0.01 70 ± 0.03

ed
Da2CS2

Da2CS3 0.57 ± 0.15 0.67 ± 0.01 1.19 ± 0.01 14.92 ± 0.01 22.74 ± 0.08 72 ± 0.09
pt
All the data were represented as average of triplicate analysis ± standard deviation.
357
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 t
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358 Table 3 Swelling and Solubility power of carboxymethylated Dioscorea starches (Da1 and Da2)

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Samples Swelling Power (%) Solubility Power (%)

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60 °C 70 °C 80 °C 90 °C 60 °C 70 °C 80 °C 90 °C

Da1CS1 6.20 ± 0.02 10.41 ± 0.01 15.25 ± 0.01 22.21 ± 0.03 4.12 ± 0.03 5.25± 0.02 9.89 ± 0.03 11.70 ± 0.08

an
Da1CS2 7.01 ± 0.07 11.21 ± 0.01 15.84 ± 0.04 22.56 ± 0.03 4.89 ± 0.04 5.62 ± 0.01 10.01± 0.02 12.05 ± 0.04

Da1CS3 7.89 ± 0.12 11.83 ± 0.08 16.01 ± 0.08 23.21 ± 0.08 5.01 ± 0.01 5.89 ± 0.01 11.20 ± 0.02 12.98 ± 0.03

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Da2CS1 6.28 ± 0.04 1.25± 0.02 16.14 ± 0.11 21.98 ± 0.01 4.15 0.07 5.65 ± 0.01 7.21 ± 0.07 9.89 ± 0.02

Da2CS2 6.98 ± 0.08 12.75 ± 0.01

ed 16.64 ± 0.02 22.67 ± 0.18 4.71 ± 0.01 6.01 ± 0.08 7.81 ± 0.11 10.12 ± 0.01
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Da2CS3 7.15 ± 0.04 13.05 ± 0.07 17.12 ± 0.04 23.77 ± 0.03 5.18 ± 0.02 6.85 ± 0.07 8.12 ± 0.03 11.06 ± 0.07
All the data were represented as average of triplicate analysis ± standard deviation.
359
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360 Table 4 Micromeritics properties of granules and physical tests of tablets prepared using carboxymethylated Dioscorea

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361 starches (Da1 and Da2)

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Samples Granule properties Physical tests of Tablets

BD (g/cm3) TD (g/cm3) HR CI (%) AR (°) Weight Thickness Diameter Friability Hardness Elastic Drug

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variation (mm) (mm) (%) (kg/cm2) recovery content (%)

(mg) (%)

M
Da1CS1 0.49 ± 0.08 0.56 ± 0.17 1.14 ± 12.50 ± 22.21 ± 0.15 249.42 ± 0.01 4.43 ± 0.01 8.00 ± 0.01 0.47 ± 0.01 4.2 ± 0.00 0.68 ± 0.00 99.24 ± 0.01

0.18 0.13

0.52 ± 0.02 0.62 ± 0.13 1.19 ± 16.12 ± 23.05± 0.08 248.83 ± 0.01 4.51 ± 0.08 8.01 ± 0.08 0.58 ± 0.01 4.3 ± 0.08 0.44 ± 0.02 99.31 ± 0.08

ed
Da1CS2

0.02 0.02

Da1CS3 0.55 ± 0.02 0.64± 0.08 1.16 ± 14.02 ± 24.25 ± 0.01 251.02 ± 0.01 4.48 ± 0.00 8.00 ± 0.00 0.32 ± 0.01 4.6 ± 0.01 0.22 ± 0.01 98.76 ± 0.01
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0.21 0.13
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Da2CS1 0.48 ± 0.08 0.55 ± 0.02 1.14 ± 12.72 ± 20.12 ± 0.08 249.81 ± 0.08 4.45 ± 0.08 8.00 ± 0.02 0.65 ± 0.08 4.1 ± 0.02 0.67 ± 0.08 100.21± 0.01

0.02 0.02

0.49 ± 0.02 0.59 ± 0.02 1.20 16.94 ± 21.17 ± 0.02 250.06 ± 0.02 4.44 ± 0.02 8.00 ± 0.02 0.53 ± 0.02 4.3 ± 0.00 0.23 ± 0.00 98.25 ± 0.08
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Da2CS2

0.13

Da2CS3 0.52 ± 0.12 0.60± 0.08 1.15 ± 13.33 ± 25.16 ± 0.01 249.52 ± 0.02 4.47 ± 0.00 8.01 ± 0.01 0.50 ± 0.01 4.5 ± 0.02 0.46 ± 0.02 100.04 ±

0.02 0.02 0.01

All the data were represented as average of triplicate analysis ± standard deviation. BD- bulk density, TD- tap density, HR- Hausner’s ratio, CI- Carr’s index, AR-angle of repose
362

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363 Figure Captions

364 Fig.1 Scanning electron micrographs of carboxymethyl derivatives of Dioscorea starches Da1

365 and Da2; A) Da1CS1 B) Da1CS2 C) Da1CS3; D) Da2CS1 E) Da2CS2 F) Da2CS3

366 Fig. 2 FTIR studies of derivatives of Dioscorea starches A) Da1 Derivatives B) Da2 Derivatives.

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367 Fig. 3 Thermogravimetry analysis (TGA) of carboxymethyl derivatives of Dioscorea starches

368 A) Da1 Derivatives-(a) Native starch, (b) Da1CS1, (c) Da1CS2, (d) Da1CS3; B) Da2 Derivatives-

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369 (a) Native starch, (b) Da2CS1, (c) Da2CS2, (d) Da2CS3

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370 Fig. 4 Release profile of carboxymethyl derivatives of Dioscorea starches.

371 an
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371

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396
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