SCIFED · saturated solution. The thermodynamic activity of the pure solute measures the escaping...

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Volume 1 · Issue 3 · 1000010SF Pharma J

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SciFed Journal of Pharmaceutics Journal

Himankar Baishya, SF Pharma J, 2018, 1:3

Selection of Co-Solvent System for Injectable Dosage Form of an Antihyperparathyroid Drug using Statistical Design

*Himankar Baishya, Manas Boxi, Zhang Zibin

*Beijing Sciecure Pharmaceuticals, Zhongbei Industrial Park, Beishicao Town, Shunyi District, Beijing, P.R. China- 101301

Keywords Paricalcitol Injection; Co-Solvent; Experimental Design; Drug Stability

Introduction Out of newly discovered drugs more than 40% drugs are lipophilic and out of which up to 40% of pharmacologically active new molecules failed to reach market only due to little or no water solubility; a serious challenge for the successful development and commercialization of new drugs in the pharmaceutical industry [1]. Therefore various formulation strategies have been investigated to improve the solubility of lipophilic drugs which includes the use of co-solvents, surfactants, lipids, micronisation, salt formation, complexation using cyclodextrins, nanoparticulate formulation, solid dispersions, etc. The solubility of a compound depends upon the physical and chemical properties of the solute and solvent as well as on such factors as temperature, pressure, the pH of the solution, and also on the state of subdivision of the solute.

The oral route remains the preferred route of drug administration due to its convenience, good patient compliance and low medicine production costs. However, many drug molecules need to be formulated in other routes due to either high degradation of the active moiety in the gastric environment, very poor absorption profile in the git, poor in-vitro-in-vivo correlation, faster onset of action

*Corresponding author: Himankar Baishya, Beijing Sciecure Pharmaceuticals, Zhongbei Industrial Park, Beishicao Town, Shunyi District, Beijing, P.R. China- 101301. E-mail: [email protected]

Received July 11, 2018; Accepted August 05, 2018; Published August 17, 2018

Citation: Himankar Baishya (2018) Selection of Co-Solvent System for Injectable Dosage Form of an Antihyperparathyroid Drug using Statistical Design. SF Pharma J 1:3. Copyright: © 2018 Himankar Baishya. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

Abstract The objective of this work was to evaluate the potential of various co-solvents for development of an injectable dosage form of Paricalcitol in a convenient as well as industrially feasible method. Paricalcitol is a potent molecule for prevention and treatment of secondary hyperparathyroidism associated with chronic kidney disease. It is marketed by Abbott Laboratories as Zemplar® in capsule as well as injectable dosage forms. We developed an injectable formulation of the same for intravenous drug delivery. pH solubility study of Paricalcitol reveals that it is insoluble at all pH values. Hence, co-solvency approach was undertaken for preparing a thermodynamically stable system with enhanced solubility. The prepared products were then analyzed for chemical stability. Experimental design was applied for selection of optimum co-solvent system as well as for prediction of the most desirable formula. The formulation design space is constructed in terms of drug stability in solution state and assay during long term and accelerated storage conditions. The selected formulation exhibited sufficient chemical stability upon storage for up to 6 months with no visible precipitation and acceptable potency.

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Citation: Himankar Baishya (2018) Selection of Co-Solvent System for Injectable Dosage Form of an Antihyperparathyroid Drug using Statistical Design. SF Pharma J 1:3.

needed or a combination of the above factors. Currently only 8% of new drug candidates have both high solubility and permeability [2]. Thus, one of the major challenges to drug development today is poor solubility, as an estimated 40% of all newly developed drugs are poorly soluble or insoluble in water. The solubility of a solute in a solvent is equal to the thermodynamic activity of the solute, usually in the pure state, divided by the activity coefficient of the solute in the saturated solution. The thermodynamic activity of the pure solute measures the escaping tendency of the solute from the pure state and decreases with increasing strengths of the solute-solute interactions, which for solid, is reflected in the melting point and lattice energy. Low solubility’s of many polar solids often results from their high melting points and high lattice energies. Modification of solid state such as the formation of amorphous forms, metastable polymorphs, co-crystals, or novel particles, cans abolish or lower the lattice energy and can therefore increase the solubility of a compound. The activity coefficient of the solute in the saturated solutions depends on the relative strengths (enthalpy) of the solute-solvent and solvent-solvent interactions and the degree of disordering (entropy) of the solvent molecules around the solute molecules [4]. The low solubility of many non-polar compounds in water result from the unfavourable negative entropy due to hydrocarbons group, known as the hydrophobic interaction. Many methods of solubility enhancement depend on the lowering the activity coefficient by means of ingenious strategies for enhancing the interactions in solutions [5]. Management of secondary hyperparathyroid has included the use of active vitamin D or vitamin D analogs for the suppression of parathyroid hormone (PTH) secretion. Although, these agents are effective, therapy is frequently limited by hypercalcemia, hyperphosphatemia, and/or eleva- vitamin D3), hypocalcemia, and hyperphosphatemia and/or elevations in the calcium-phosphorus (Ca-P) product [6]. Both oral and parenteral forms of calcitriol have been effective in treating and preventing secondary hyperparathyroidism. However, intravenous calcitriol has been more widely used and thought to be the more effective form [7]. We intend to develop an intravenous formulation of Paricalcitol for clinical management of secondary hyperparathyroidism.

Materials and Methods Due to the hydrophobic nature of Paricalcitol, the

main target during drug development stage was selection of appropriate parenteral excipients within acceptable limits for complete solubilization of the API and the resultant solution remain stable during storage during its shelf life. Co-solvency approach was undertaken so as to increase the solubility of Paricalcitol. Two critical parameters associated with drug solubility were identified:(i) Chemical stability of drug product in aqueous media(ii) Precipitation of Paricalcitol

The obtained formulations were subjected to initial Freeze-Thaw studies for screening of appropriate co-solvent systems for Paricalcitol followed by stability studies at accelerated as well as normal conditions. Formulation development was started by using a co-solvent with similar properties as propylene glycol. Ethanol as the co-solvent was used in accordance with innovator product. In the initial trials, Glycerol 99.5% was used as the solubilizer. Furthur trials were taken using PEG 200, Polysorbate 80, PEG 300, PEG 400 and Macrogol esters. Statistical experimental design was used to obtain the co-solvent system with maximum desirability for the critical parameters. The formulation design space was constructed in terms of variability in concentration of co-solvent. (Table 1) Freeze Thaw study was carried out by subjecting the samples to alternating environments of 2-8ºC and 40ºC every 7 days. Samples were observed for presence / absence of precipitation of Paricalcitol or any other component of the formulation.

Statistical Analysis The various parameters analyzed were subjected to statistical analysis using Design Expert Pro (Version 10). Statistical significance was determined by one-way ANOVA, with p<0.05 being considered significant.

Results All the formulations were subjected to Freeze Thaw study followed by stability study at accelerated and normal storage conditions.

● Precipitation was observed at 4˚C in Experiment 1 after 1 month storage during Freeze Thaw studies

● No precipitation was observed in case of Experiment 2 containing PEG 200 during 3 months of Freeze Thaw




Table 1: Composition of Paricalcitol Injection using Various Solubilizers

IngredientsComposition (% v/v)

Experiment 1 Experiment 2 Experiment 3 Experiment 4 Experiment 5 Experiment 6

Paricalcitol 0.0005 0.0005 0.0005 0.0005 0.0005 0.0005

Ethanol 20.00 20.00 20.00 20.00 20.00 20.00

Glycerol 99.5% 15.00 --- --- --- --- ---

PEG 200 --- 15.00 --- --- --- ---

Polysorbate 80 --- --- 10.00 --- --- ---

PEG 300 --- --- --- 15.00 --- ---

PEG 400 --- --- --- --- 12.00 ---

Macrogol 15 Hydroxystereate --- --- --- --- --- 0.50

WFI Q.S. Q.S. Q.S. Q.S. Q.S. Q.S.

studies. However, there occurs a steady decline in drug content during 3 months at accelerated and normal storage conditions.(Graph 1)

● Furthur trials were conducted using Polysorbate 80 as the solubilizing agent. The results obtained were similar to the results obtained using PEG 200. However, the rate of degradation during storage was less intense that that with PEG 200.(Graph 2)

● Since results of PEG 200 showed no precipitation, it was decided to study of the formulations in other molecular weight PEGs. Two other formulations containing PEG 300 and PEG 400 were prepared. ● PEG 300 samples showed good stability for 3 months at all time points. However when the samples were autoclaved initially and then loaded for stability studies, a decrease in drug content was observed, especially at 40˚C. (Graph 3, Graph 4, Graph 5)

● Final Experiment was taken with Macrogol 15 Hydroxystereate as the co-solvent. When 0.5% of Macrogol 15 Hydroxystereate was used as co-solvent, no precipitation was observed during Freeze-Thaw studies for a period of 6 months. Also, the drug product showed

satisfactory results with regard to assay at all conditions for a duration of upto 6 months.(Graph 6 Graph 7, Figure 1) The above phase diagram was constructed to ascertain the degree of solubilization of Paricalcitol in PEG 300, Glycerol and Macrogol 15 Hydroxystereate. It was observed that at 0.7% concentration of co-solvent system, both PEG 300 and glycerol 99.5% show solubility of about 11% whereas 100% solubility is achieved at around 0.4% Macrogol 15 Hydroxystereate. Macrogol 15 Hydroxystereate is found to be the most effective solubilizer for Paricalcitol. (Figure 2) The above phase diagram explains the role of different molecular weight PEG with respect to the rate of solubilization of Paricalcitol using 20% Ethanol as the co-solvent. PEG 200 shows a slightly better solubility profile as compared to PEG 300. However, formulations made using PEG 300 are comparatively more stable during storage especially at a temperature of 4⁰C. (Figure 3) All PEG grades shows a similar solubility profile irrespective of their molecular weight. However, formulations made using PEG 300 are comparatively more stable during storage especially at a temperature of 4⁰C. (Figure 4)




Graph 2: Freeze Thaw stability of Paricalcitol Injection with Polysorbate 80 as Solubilizer

Graph 1: Freeze Thaw Stability of Paricalcitol Injection with PEG 200 as Solubilizer

Graph 3: Freeze Thaw stability of Paricalcitol Injection with PEG 300 as Solubilizer




Graph 4: Freeze Thaw stability of Paricalcitol Injection with PEG 300 as solubilizer (after Autoclaving at 121ºC for 15 Minutes)

Graph 5: Freeze Thaw stability of Paricalcitol Injection with PEG 400 as Solubilizer

Graph 6: Freeze Thaw stability of Paricalcitol Injection with Macrogol 15 Hydroxystereate as Solubilizer




Graph 7: Freeze Thaw stability of Paricalcitol Injection with Macrogol 15 Hydroxystereate as Solubilizer (after Autoclaving at 121ºC for 15 Minutes)

Design-Expert® Software Component Coding: Actual % Dissolved (%)

100 11.2

X1 = G: MACROGOL 15 HYDROXYSTEREATE X2 = D: PEG 300 X3 = B: Glycerol 99.5% Actual Components A: Paricalcitol = 0.0005 C: PEG 200 = 0 E: PEG 400 = 0 F: POLYSORBATE 80 = 0 H: WFI = 79.2828 J: Ethanol = 20

G: MACROGOL 15 HS (%V/V) 0.716667

D: PEG 300 (%V/V) 0.716667 B: Glycerol 99.5% (%V/V) 0.716667

0 0

0 % Dissolved (%)

20

40

60

80

100

Figure 1: Comparative Solubility of Paricalcitol: Macrogol 15 HS vs PEG 300 vs Glycerol


100 11.2

X1 = H: WFI X2 = C: PEG 200 X3 = D: PEG 300 Actual Components A: Paricalcitol = 0.0005 B: Glycerol 99.5% = 0 E: PEG 400 = 0 F: POLYSORBATE 80 = 1.74728 G: MACROGOL 15 HYDROXYSTEREATE = 0 J: Ethanol = 20

H: WFI (%V/V) 78.2522

C: PEG 200 (%V/V) 13.2527

D: PEG 300 (%V/V) 13.2527

0 0

64.9995

% Dissolved (%)

40

60

80

100

Figure 2: Comparative Solubility of Paricalcitol: PEG 200 vs PEG 300




Figure 3: Similar Dissolving Capabilities of PEG 200, PEG 300 and PEG 400


100 11.2

X1 = G: MACROGOL 15 HYDROXYSTEREATE X2 = F: POLYSORBATE 80 X3 = H: WFI Actual Components A: Paricalcitol = 0.0005 B: Glycerol 99.5% = 0 C: PEG 200 = 0 D: PEG 300 = 0 E: PEG 400 = 0 J: Ethanol = 20

% Dissolved (%)

G: MACROGOL 15 HS (%V/V) 0.5

F: POLYSORBATE 80 (%V/V) 8.9286 H: WFI (%V/V) 79.8995

64.9995 0

0

20

40

60

80

100

Figure 4: Comparative solubility of Paricalcitol in Various Solvents (Macrogol 15 Hydroxystereate vs Polysorbate 80)

The above contour plot studies the degree of solubility of Paricalcitol using a system consisting of Polysorbate (0-10%), Macrogol 15 Hydroxystereate (0-0.5%) and WFI (65-79.9%) keeping concentration of Paricalcitol and Ethanol constant at 0.0005% and 20% respectively. It was observed that the solubility increased as a function of co-solvent concentration. The resultant co-solvent system showed desirability of around 60% with Polysorbate 80 as the co-solvent. However, the desirability increases to 100% upon incorporation of Macrogol 15 Hydroxystereate into the co-solvent system. (Table 2, Figure 5) The above 3D contour plot studies the degree of solubility of Paricalcitol using similar concentrations of PEG 200, Macrogol 15 Hydroxystereate and PEG 300 at concentration range of 0-0.7 % for each component keeping concentration of Paricalcitol , Ethanol and WFI constant at 0.0005%, 20% and 79.3% respectively. It

was observed that even after using the maximum amount of PEG 200 and PEG 300 (0.75% respectively) the desirability of the system is very low at 11.2%. However, upon incorporation of Macrogol 15 Hydroxystereate into the system, the solubility and desirability of the system was seen to increase in a dose dependent manner. (Figure 6) The above 3D contour plot studies the degree of solubility of Paricalcitol using similar concentrations of PEG 400, Macrogol 15 Hydroxystereate and Polysorbate 80 at concentration range of 0-0.75 % for each component keeping concentration of Paricalcitol , Ethanol and WFI constant at 0.0005%, 20% and 79.3% respectively. The results obtained were in-line to the ones obtained in the previous contour mapping thereby confirming that Macrogol 15 Hydroxystereate provides the best co-solvent system for ethanol for the solubilization of Paricalcitol. (Table 3, Table 4, Figure 7)




Table 2: Change-Response Matrix of Paricalcitol Injection

Name Units Type Change Low High

Paricalcitol %V/V Component Hard 0.0005 0.0005Glycerol 99.5% %V/V Component Easy 0 15

PEG 200 %V/V Component Easy 0 15PEG 300 %V/V Component Easy 0 15PEG 400 %V/V Component Easy 0 12

POLYSORBATE 80 %V/V Component Easy 0 10MACROGOL 15

HYDROXYSTEREATE %V/V Component Easy 0 0.5

WFI %V/V Component Easy 64.9995 79.8995Ethanol %V/V Component Hard 20 20

% Dissolved % Response 11.2 100Freeze Thaw Months Response 0 6

Figure 5: Comparative Solubility: Macrogol 15 Hydroxystereate vs PEG 200 vs PEG 300

Figure 6: Comparative Solubility: Macrogol 15 Hydroxystereate vs PEG 400 vs Polysorbate 80




Table 3: ANOVA (% Solubilized) [Partial Sum of Squares - Type III]

Source Sum of Squares df Mean Square F Value p-value Prob > F SignificanceModel 27413.80 6 4568.97 42276.08 < 0.0001 Significant

Linear Mixture 27413.80 6 4568.97 42276.08 < 0.0001Residual 2.49 23 0.11Cor Total 27416.29 29

The Model F-value of 42276.08 implies the model is significant. There is only a 0.01% chance that an F-value this large could occur due to noise.Values of “Prob > F” less than 0.0500 indicate model terms are significant.In this case B, C, D, E, F, G are significant model terms.

Table 4: Degree of Solubilization of Paricalcitol using Mathematical Model

Final Equation in Terms of Real Components:% Dissolved =+668.41973 * Glycerol 99.5%+747.17547 * PEG 200+667.28119 * PEG 300+1120.45597 * PEG 400+1120.61594 * POLYSORBATE 80

+20046.92275 * MACROGOL 15 HYDROXYSTEREATE-0.048619 * WFI

Final Equation in Terms of Actual Components:% Dissolved =

+6.68420 * Glycerol 99.5%+7.47175 * PEG 200+6.67281 * PEG 300+11.20456 * PEG 400+11.20616 * POLYSORBATE 80

+200.46923 * MACROGOL 15 HYDROXYSTEREATE-4.86191E-004 * WFI

Design-Expert® Software Component Coding: Actual Freeze Thaw (Months)

6 0

X1 = G: MACROGOL 15 HYDROXYSTEREATE X2 = C: PEG 200 X3 = D: PEG 300 Actual Components A: Paricalcitol = 0.0005 B: Glycerol 99.5% = 2.93542 E: PEG 400 = 1.74728 F: POLYSORBATE 80 = 1.74728 H: WFI = 67.9153 J: Ethanol = 20

Freeze Thaw (Months)

G: MACROGOL 15 HYDROXYSTEREATE (%V/V) 0.5

C: PEG 200 (%V/V) 5.33638

D: PEG 300 (%V/V) 5.65418

0 0.232356

0

2

3

4

5

Figure 7: Effect of Solvents on Freeze - Thaw Stability Solutions




Freeze-thaw studies were then undertaken to study the stability of the respective co-solvent systems using 20% ethanol as the primary solvent. Macrogol 15 Hydroxystereate (0-0.5%), PEG 300 (0- 5.5%) and PEG 200 (0-5.3%) were used as the co-solvent components along with constant concentrations of Ethanol (20%), Paricalcitol (0.0005%), Glycerol (2.93%), Polysorbate 80 (1.75%), PEG 400 (1.75%) and WFI (67.9%). Samples were kept at 25ºC for 1 week followed by 2-8ºC for 1 week and studies for any crystal growth or precipitation. The cysle was repeated until any precipitation was observed in the samples. PEG 200 and PEG 300 showed stablility of 1 months each while the stability of the system increased to 6 months Macrogol 15 Hydroxystereate in the co-solvent system.(Figure 8a) Macrogol 15 Hydroxystereate (0-0.5%), Polysorbate (0- 3,6%) and PEG 400 (0-3.6%) were used as the co-solvent components along with constant concentrations of Ethanol (20%), Paricalcitol (0.0005%), Glycerol (2.93%), PEG 200(2.8%), PEG 300 (2.9%) and

WFI (67.9%). Samples were kept at 25ºC for 1 week followed by 2-8ºC for 1 week and studies for any crystal growth or precipitation. The cycle was repeated until any precipitation was observed in the samples. Polysorbate, PEG 300 and Macrogol 15 Hydroxystereate showed stablility of 3 months, 2 months and 6 months respectively.(Figure 8b) Polysorbate 80 (0-11.2%), Glycerol (0-11.2%) and PEG 400 (0-11.2%) were used as the co-solvent components along with constant concentrations of Ethanol (20%), Paricalcitol (0.0005%), Glycerol (2.93%), PEG 300 (3.78%) and WFI (65%). PEG 200 and Macrogol 15 Hydroxystereate were not considered in the study. Samples were kept at 25ºC for 1 week followed by 2-8ºC for 1 week and studies for any crystal growth or precipitation. The cycle was repeated until any precipitation was observed in the samples. Glycerol showed a maximum duration of 1 month stability whereas solution contaioning Polysorbate 80 or PEG 400 are stable for 3 months. (Figure 8c)

Design-Expert® Software Component Coding: Actual Freeze Thaw (Months)

6 0

X1 = B: Glycerol 99.5% X2 = F: POLYSORBATE 80 X3 = E: PEG 400 Actual Components A: Paricalcitol = 0.0005 C: PEG 200 = 0 D: PEG 300 = 3.77973 G: MACROGOL 15 HYDROXYSTEREATE = 0 H: WFI = 64.9995 J: Ethanol = 20

B: Glycerol 99.5% (%V/V) 11.2203

F: POLYSORBATE 80 (%V/V) 11.2203

E: PEG 400 (%V/V) 11.2203

0 0

0

Freeze Thaw (Months)

1.5

2

2.5

3

Figure 8a: Effect of solvents on Freeze - Thaw Stability Solutions

Figure 8b: Comparative Freeze-Thaw Data for Glycerol vs Polysorbate 80 vs PEG 400




Acknowledgement We would like to thank all the colleagues of Formulation Development Department of Beijing Sciecure Pharmaceuticals for assisting in our work and providing their valuable inputs for the successful completion of this project.

Conclusion A thermodynamically stable injection dosage form of Paricalcitol has been developed using co-solvency approach in a cost effective and industrially feasible process. Statistical design methodology was employed to ascertain the most desirable formulation for injectable formulation of Paricalcitol considering the rate of solubilization of Paricalcitol and the stability of the dosage forms to freeze-thaw cycling as the two primary parameters for study. Macrogol esters offer the most desirable solubility for Paricalcitol along with good physicochemical stability of the obtained product at real time and accelerated conditions. By use of experimental design, an optimized formulation was obtained with maximum desirability (% Dissolved and Duration of Freeze thaw stability as response parameters).

References1. Sinko PJ (2006) Martin’s physical pharmacy and pharmaceutical sciences. 5th edition, 231-264.

2. Naseem A, Olliff CJ, Martini LG, et al. (2004) Effects of plasma irradiation on the wettability and dissolution of compacts of griseofulvin. ‎Int J Phar 269: 443-450.

3. Finholt P, Solvang S (1968) Dissolution kinetics of drugs in human gastric juice the role of surface tension. J Pharm Sci 57: 1322-1326.

4. Pinnamaneni S, Das NG, Das SK (2002) Formulation approaches for orally administered poorly soluble drugs. Dei Pharmazie 57: 291-300.

5. Lawrence MJ, Rees GJ (2000) Microemulsion‐based media as novel drug delivery systems. Adv Drug Deliv Rev 45: 89–121.

6. Sprague SM, Llach F, Amdahl M, et al. (2003) Paricalcitol versus calcitriol in the treatment of secondary hyperparathyroidism. Kidney International 63: 1483-1490.

7. Slatopolsky E, Weerts C, Thielan J (1984) Marked suppression of secondary hyperparathyroidism by intravenous administration of 1,25-dihydroxy-cholecalciferol in uremic patients. J Clin Invest 74: 2136-2143.

Design-Expert® Software Component Coding: Actual Desirability

1.000 0.000

X1 = D: PEG 300 X2 = E: PEG 400 X3 = G: MACROGOL 15 HYDROXYSTEREATE Actual Components A: Paricalcitol = 0.0005 B: Glycerol 99.5% = 0 C: PEG 200 = 2.90196 F: POLYSORBATE 80 = 1.17886 H: WFI = 75.4384 J: Ethanol = 20

Desirability

D: PEG 300 (%V/V) 0.480321

E: PEG 400 (%V/V) 0.480321 G: MACROGOL 15 HYDROXYSTEREATE (%V/V) 0.480321

0 0

0

0.2 0.4

0.6

0.8 Prediction 0.870

Figure 8c: Optimized Formulation as Per Design Expert Pro with Maximum Desirability (% Dissolved and Duration of Freeze thaw Stability as Response Parameters)


SCIFED · saturated solution. The thermodynamic activity of the pure solute measures the escaping...

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