1. INTRODUCTION:

Zopiclone is a non-benzodiazepine sedative hypnotic drug used for the short-term treatment of insomnia. It is a (atypical antipsychotic) psychotropic agent that belongs to the thienobenzodiazepine class¹. Immediate release zopiclone tablets are available in the market for oral administration. Multiparticulate drug delivery system have various advantages over single-unit dosage forms because of their potential benefits like flexible release patterns, predictable gastric emptying, no risk of dose dumping and increased bioavailability with less inter and intra-subject variability².

Pellets are the agglomerates obtained from diverse starting materials of granules or fine powders of excipients and bulk drugs utilizing different pelletization techniques^3,4.

The size range of pellets is about 0.5 to 2.0mm. Various pelletization techniques reported in literature are, balling, compression, extrusion/spheronization, solution/suspension layering and spray drying/ congelling.

Pellets consist of various formulation aids such as filler/diluents, (lactose, microcrystalline cellulose, starch) binders, (hydroxyl propylmethyl cellulose, polyvinylpyrrolidone), lubricants, (magnesium stearate), separating agent, (Talc), disintegrants, (croscarmellose sodium, sodium starch glycolate) and spheronization enhancer, (microcrystalline cellulose)⁵. Umalkar et. al. have reported development of mouth dissolving tablet of zopiclone⁶. Swapna and Jithan have reported sustained release microspheres of zopiclone for parenteral use⁷.

The objective of this study was to formulate zopiclone immediate release pellets. For this purpose, MCC, SSG, HPMC E-5, corn starch and lactose were used. Concentrations of SSG and HPMC E-5were optimized using 3²factorial design.

2. MATERIALS AND METHODS:

2.1 Materials:

Zopiclone was obtained as a gift sample from Calyx Chemicals and Pharmaceuticals Ltd., Tarapur, Boisar. Microcrystalline cellulose (Avicel PH101) was purchased from Research Lab, Mumbai. Corn starch, sodium starch glycolate, HPMC E-5, lactose (monohydrate), isopropyl alcohol and hydrochloric acid were purchased from Loba Chemie, Mumbai. Polyvinylpyrrolidone (PVP) K-30 was purchased from Himedia Laboratories Pvt. Ltd., Mumbai.

2.2 Methods:

2.2.1 Preparation of calibration curve in 0.1N HCl:

Zopiclone 10mg was weighed accurately and transferred to 100ml volumetric flask. Methanol (20ml) was added and sonicated for 15 min to dissolve the drug. Volume was made up to 100ml with 0.1N HCl. Further concentrations were made (2.5, 5, 10, 15, 20µg/ml) from stock solution by appropriate dilution using 0.1N HCl. The solutions were analysed by UV visible spectrophotometer (Shimadzu) at 305nm, using the 0.1N HCl as a blank. Calibration curve was plotted. Similarly, calibration curve was also prepared in methanol.

2.2.2 Drug excipients compatibility studies:

During formulation of pellets there are chances of drug degradation as well as unstable product formation due to drug excipient interactions. Hence drug excipients compatibility study is required. For this study drug was mixed with different excipients in ratio (1:1). These mixtures were kept in dry box for making it moisture free. Then they were mixed separately with potassium bromide with trituration. Each mixture was placed in DRS assembly sample holder. The infrared spectra of all mixtures (excipient + drug) were recorded using shimadzu FTIR spectrophotometer (Shimadzu, 8400S, Japan) over the range of 4000-400 cm^-1with a resolution of 5 cm^-1and observed for the interaction between drug and excipients.

2.2.3. Preparation of Pellets:

Pellet formulations were prepared by changing the amounts of corn starch, HPMC E-5, and sodium starch glycolate (SSG), using extrusion–spheronization method. MCC was used as pelletizing agent. HPMC E-5 was used as a binder. Lactose and corn starch were used as ﬁllers and SSG was used as a disintegrant.

Pellets were formulated by various steps using lab scale spheronizer.

2.2.3.1. Preparation of wet mass:

The powder mixture was prepared by homogenous blending of drug, microcrystalline cellulose, corn starch, HPMC E-5, lactose and sodium starch glycolate. Binder solvent containing purified water and isopropyl alcohol (8:2) was added to dry mixture. Consecutive pauses were taken during the addition of the liquid binders for proper kneading to obtain a damp mass of appropriate consistency for the extrusion process⁸.

2.2.3.2. Extrusion:

Prepared wet mass was immediately extruded through sieve number 16 (B.S.S.) to obtain the rod shape extrudates.

2.2.3.3. Spheronization:

The extrudates were spheronised at 850rpm with 5mm friction plate to obtain spherical pellets. Wet pellets were dried at room temperature.Fig.1 shows photograph of typical pellets.

Fig.1: Prepared pellets

Table 1: Composition of zopiclone pellets

Batch code	MCC (g)	SSG (g)	HPMCE-5 (g)	Corn starch (g)	Lactose (g)	Drug (g)	Total (g)
F1	6.4	1.6 (8%)	0.4 (2%)	10.8	0.425	0.375	20
F2	6.4	1.6 (8%)	0.8 (4%)	10.4	0.425	0.375	20
F3	6.4	1.6 (8%)	1.2 (6%)	10	0.425	0.375	20
F4	6.4	1.2 (6%)	0.4 (2%)	11.2	0.425	0.375	20
F5	6.4	1.2 (6%)	0.8 (4%)	10.8	0.425	0.375	20
F6	6.4	1.2 (6%)	1.2 (6%)	10.4	0.425	0.375	20
F7	6.4	0.8 (4%)	0.4 (2%)	11.6	0.425	0.375	20
F8	6.4	0.8 (4%)	0.8 (4%)	11.2	0.425	0.375	20
F9	6.4	0.8 (4%)	1.2 (6%)	10.8	0.425	0.375	20
F10	6.4	1.2 (6%)	0.8 (4%)	10.8	0.425	0.375	20
F11	6.4	1.2 (6%)	0.8 (4%)	10.8	0.425	0.375	20
F12	6.4	1.2 (6%)	0.8 (4%)	10.8	0.425	0.375	20
F13	6.4	1.2 (6%)	0.8 (4%)	10.8	0.425	0.375	20

2.2.4 Formula optimization:

Preliminary studies showed a significant effect of concentration of SSG and HPMCE-5 on particle size, Hardness, disintegration time and percent drug release from pellets. These two parameters were selected as independent variables with three levels resulting in 13 experimental runs (Table 1).

SSG concentration (A) was at levels 4, 6, 8% w/w and HPMC E-5concentration (B) was at levels 2, 4, 6% w/w. The effects of these variables on responses, disintegration time (Y1), drug release after 45 min (Y2), practical yield (Y₃), pellets size (Y₄), hardness (Y₅), bulk density (Y₆) and tapped density (Y₇) were evaluated. The resulting data was fitted into Design Expert software (Design Expert, version 11) (Stat-ease, USA). The statistical significance of the data was evaluated using analysis of variance (ANOVA) The surface response plots were produced to study the collaborative effects of the variables..

2.2.5 Evaluation of pellets:

2.2.5.1 Determination of % practical Yield:

The percentage yield of pellets was calculated by the following formula:

Practical yield of pellets (gm)

% Pactical yield= ––––––––––––––––––––––––––×100

Amount of powder mixture (gm)

2.2.5.2 Bulk Density:

Bulk density was determined by taking weighed amount of pellets into 50ml graduated cylinder and volume was measured. Bulk density was calculated by using following formula:

Mass

Bulk density = ––––––––––––

Bulk Volume

2.2.5.3 Tapped Density:

Pellets were filled in 50ml graduated cylinder and tapped until the powder bed volume reached a minimum. Tapped density was calculated by formula:

Mass

Tapped Density = ––––––––––––––––

Tapped volume

2.2.5.4 Hausner’s ratio:

Hausner’s ratio gives an idea regarding the flow of the pellets. Hausner’s ratio was calculated by following equation:

Tapped density

Hausner’s ratio= –––––––––––––––

Bulk density

2.2.5.5 Carr’s Index:

The Carr’s index was calculated by comparing bulk density and tapped density of the pellets using the following formula:

Tapped density – Bulk density

Carr’s index= ––––––––––––––––––––––––––––×100

Tapped density

2.2.5.6 Angle of repose:

Angle of repose was determined by using fixed funnel method. The angle of repose was calculated from the height (h) and average radius(r) of powder cone:

Angle of repose (θ) = tan^-1(h/r)

2.2.5.7 Particle size (pellets size):

The particle size of prepared pellets was measured using digital vernier caliper. The mean diameter of 20 pellets in each batch was noted as particle size.

2.2.5.8 Hardness:

Hardness of pellets was measured by using a digital pellets hardness tester. Hardness of 6 pellets of each batch was measured.

2.2.5.9 Disintegration time:

Disintegration time of pellets from each batch was determined by noting the disintegration time in distilled water. There is no standard disintegration apparatus for disintegration of pellets. Accurately weighed 400mg pellets were put in 50ml distilled water and disintegration time was noted down by shaking them in water bath shaker at 37°C till disintegration.

2.2.5.10 Drug content:

Pellets were crushed in a mortar to obtain fine powder. 400mg powder was accurately weighed and dissolved in 100ml methanol and sonicated for 5 min. Then resulting solution was filtered using whatman filter paper, one ml of filterate was diluted to 10ml by using methanol. The resulting solution was measured at 305 nm using UV-Spectrophotometer. Concentration of drug solution was calculated by using equation of line showed in calibration curve in methanol.

Actual amount of zopiclone found in pellets

% Pactical yield= ––––––––––––––––––––––––––––––––––––– ×100

Theoretical amount of zopiclone in pellets

2.2.5.11 In-vitro dissolution studies:

In vitro dissolution studies were performed by using IP dissolution procedure given for zopiclone tablets⁹. Accurately weighed sample of pellets containing 7.5mg of zopiclone was dropped into 500ml of 0.1N HCl maintained at the temperature 37°C ± 0.5°C. The paddle was rotated at a speed of 50 RPM. At different time intervals, 10ml aliquot of the sample was withdrawn and the volume was replaced with fresh dissolution medium.

The collected samples were filtered with whatman filter paper, filtrate was mixed and absorbance of filtrate was measured at 305 nm. Drug release was calculated using standard curve in 0.1 N HCl.

Table 2: Calibration curve equations and multiple correlation coefficients of zopiclone.

Sr. No	Solvent	Equation	R²
1	0.1 N HCl	Y= 0.0372x+0.00790	0.999
2	Methanol	Y= 0.0411x-0.00254	0.992

3. RESULTS AND DISCUSSION:

3.1 Calibration curve of Zopiclone

Calibration curve of zopiclone in 0.1 N HCL and methanol were linear in range of 2.5 µg/ml to 20 µg/ ml.

3.2 Excipient compatibility study:

Drug-excipient compatibility for zopiclone along with the excipients used in the formulation was checked using FTIR spectroscopy. As shown in fig.2a, carbonyl stretching of the zopiclone in IR was found at 1722.49 cm^-1. However, while in mixture of zopiclone with PVP K-30, it was found to shift from 1722.49 to 1701.27 cm^-1 which showed strong interaction of zopiclone with PVP K-30 (Fig.2c). Since PVP K-30 was incompatible with zopiclone. HPMC E-5 was selected as binder for carrying out the formulation. Zopiclone with HPMC E-5 showed carbonyl stretching at 1718.63 cm^-1which indicated no interaction with HPMC E-5(Fig.2d). Other excipients did not show interaction with zopiclone.

Fig.2: FTIR spectra of (a) zopiclone (b) PVP K-30 (c) physical mixture of zopiclone and PVP K-30 (d) physical mixture zopiclone and HPMC E-5.

Table 3: Evaluation of zopiclone pellets

Batch code	Practical yield (%)	Particle size (mm)	Hardness (Kg/cm²)	Drug content (%)	B. D (gm/ml)	T. D (gm/ml)	Carrs indx (%)	Hausner ratio	Angle of repose (θ)
F1	71.01	1.08	1.6	89.24	0.750	0.789	4.94	1.05	29.85
F2	71.72	2.19	0.76	91.73	0.786	0.832	5.52	1.05	27.20
F3	82	2.06	1.33	97.12	0.747	0.792	5.68	1.06	30.37
F4	79.5	1.48	1.15	92.10	0.729	0.820	11.17	1.12	29.03
F5	81.28	1.50	1.45	92.48	0.685	0.796	13.96	1.16	31.21
F6	73.5	1.10	0.95	94.86	0.802	0.906	11.62	1.12	29.72
F7	72.25	1.26	1.75	89.75	0.725	0.773	6.27	1.06	29.37
F8	77.1	1.55	1.5	91.89	0.802	0.884	9.27	1.10	31.34
F9	81.33	1.21	1.95	93.95	0.776	0.880	11.81	1.33	29.43
F10	78.6	1.07	1.05	92.16	0.856	0.931	8.16	1.08	32.45
F11	82	1.24	1.23	93.35	0.810	0.892	9.26	1.10	34.56
F12	77.18	1.32	1.16	92.43	0.788	0.876	10.10	1.11	27.11
F13	86	1.13	1.4	93.01	0.827	0.931	11.19	1.12	28.36

3.3 Optimization of formulation:

Data of evaluation of zopiclone pellets is shown in table 3 and 4. Effect of independent variables i.e. concentration of disintegrant (A) and concentration of binder (B) was evaluated on disintegration time (Y₁), drug release at 45 min (Y₂), practical yield (Y₃), pellets size (Y₄), hardness (Y₅), bulk density (Y₆) and tapped density (Y₇) using design expert software (Design Expert, version 11) (Stat-ease, USA). Effect on disintegration time (Y₁) and % drug release (Y₂) showed significance so those are discussed here.

Table 4: Evaluation of zopiclone pellets (Responses)

Batch code	Disintegration time (min)	Drug release After 45 min(%)
F1	6.27	96.5
F2	8.06	92.5
F3	10.37	97.1
F4	6.31	97.7
F5	9.04	94.3
F6	10.37	96.4
F7	8.32	94.8
F8	8.34	89.4
F9	9.88	92.6
F10	7.03	94.1
F11	8.32	96.3
F12	7.56	98.4
F13	7.35	96.2

3.3.1. Effect on disintegration time:

The disintegration time of prepared pellets was found to be from 6.27 to10.37min. Linear model suggested by Design expert software for disintegration time is as following and is significant (P is 0.0018).

Disintegration time (Y₁) =8.25-0.31A+1.62B........... (1)

From equation (1) negative coefficient of A indicated decrease in disintegration time with increase in SSG concentration and positive coefficient of B indicated increase in disintegration time with increase in HPMC E-5 concentration. This correlates well with SSG as a disintegrant and HPMC E-5as a binder. The counterplots and 3D response plot indicating the relative effect of concentration of SSG and HPMC E-5 on the disintegration time of zopiclone pellets are shown in fig. 3A and 3B respectively.

Fig. 3A

Fig. 3B

Fig. 3: (A) Counterplot showing the effect of various levels of two independent variables, (B) 3D Response plot showing the influence of the concentration of SSG and HPMC E-5 on the Disintegration time.

3.3.2 Effect on %drug release:

The percent drug release after 45 min. is one of the important parameters to be optimized. Quadratic model suggested by design expert software for % drug release is as following and is significant (P is 0.0457)

Drug release (Y₂) =95.43+1.55A-0.48B+0.70AB-3.41A²+2.69B².. (2)

Equation (2) presented positive coefficient of A indicated increase in percent drug release after 45 min (Y₂) with increase in SSG concentration and negative coefficient of B indicated decrease in percent drug release after 45 min with increase in HPMC E-5 concentration. This also correlates with SSG as a disintegrant and HPMC E-5 as a binder. The counterplots and 3D response plot indicating the relative effect of concentration of SSG and HPMC E-5 on percent drug release after 45 min of zopiclone pellets are shown in fig. 4A and 4B respectively.

Fig. 4A

Fig. 4B

Fig. 4: (A) Counterplot showing the effect of various levels of two independent variables, (B) 3D response plot showing the effect of the concentration of SSG and HPMC E-5on the % drug release of pellets after 45 min.

From desirability search, using Design Expert software, F7 was selected as optimized formulation. In order to validate the model, F7 batch was prepared again and analyzed for disintegration time and % drug release after 45 minutes. Table 5 shows the predicted and actual values for F7 formulation.

Fig. 5: Comparative percent release profile of optimized formulation of pellet filled in a capsule with only pellets of zopiclone.

Table 5: Validation of optimised formulation (Batch code: F7)

Parameters	Predicted value	Actual (experimental) value
Disintegration time (min)	7	8.01
Drug release after 45 min(%)	94	96.7

Optimized batch of zopiclone pellets (F7) was filled into capsule shell equivalent to 7.5 mg of the drug and it’s percent release was obtained. The drug release was compared for only pellets and pellets filled in capsule as shown in Fig.5.The capsule shells did not retard drug release.(Similarity factor F2= 86.08).

4. CONCLUSION:

From excipient compatibility study, compatible excipients like SSG, microcrystalline cellulose, HPMC E-5, corn starch and lactose monohydrate were selected for pellet formulation. In preliminary study, it was found that SSG and HPMC E-5 concentrations may influence critical quality parameters like disintegration time and %drug release after 45 min. By using factorial design, the zopiclone pellets batch (F7) prepared with SSG (4%) and HPMC E-5 (2%) spheronized at 850 RPM was found to be optimized. The optimized batch shows disintegration time8.32 min and % drug release of 94.8 % after 45 min. Pellets formulated in this study can serve as an alternative to tablet dosage form.

5. ACKNOWLEDGEMENT:

Authors are thankful to Calyx Chemicals and Pharmaceuticals Ltd., Tarapur, Boisar for providing the gift sample of zopiclone. Authors are thankful to Dr. R. K. Nanda for his expert opinion on FTIR spectra.

6. REFERENCES:

1. David. J.The pharmacology and mechanisms of action of new generation, non-benzodiazepine hypnotic agents.CNS Drugs, 2004; 18 (1):9-15.

2. Shelar. V. S, Shirolkar S. V, Kale R.N. Formulation optimization of promethazine theoclate immediate release pellets by using extrusion-spheronization technique. International Journal of Applied Pharmaceutics, 2018; 10(1) :30-35.

3. Patil. S. M, Mishra, R. V, Shirolkar. S. V. Development and evaluation of sustain release simvastatin pellets. Research Journal of Pharmacy and Technology, 2017; 10(8):2017-2020.

4. Muley.S, Nandgude.T, Poddar. S. Extrusion - spheronization a promising pelletization technique: In-depth review. Asian Journal of Pharmaceutical Sciences, 2016; 2:684–699.

5. Han. X, Wang. L, Sun.Y, Liu.X, Liu.W, Du.Y, li.L, Sun.J.Preparation and evaluation of sustained-release diltiazem hydrochloride pellets. Asian Journal of Pharmaceutical Sciences, 2013; 8(4): 244–251.

6. Umalkar. D.G, Rathinaraj.B. S, Shinde. G. V.Design and evaluation of mouth dissolving tablet of zopiclone using different superdisintegrants.Journal of Pharmaceutical Sciences and Research, 2013; 2(8): 527–533.

7. Swapna. N, Jithan. Av.Preparation, characterization and in vivo evaluation of parenteral sustained release microsphere formulation of zopiclone.Journal of Young Pharmacists, 2010; 2(3): 223–228.

8. Rahman. M. A, Ahuja. A, Baboota. S, Bali. V, Saigal. N, Ali. J. Recent advances in pelletization technique for oral drug delivery: a review. Curr Drug Delivery, 2009; 6:122-129.

9. Indian Pharmacopoeia, 2014; 3:3023-3025.

Received on 30.06.2019 Modified on 03.09.2019

Research J. Pharm. and Tech 2020; 13(3):1131-1136.

DOI: 10.5958/0974-360X.2020.00208.5