LYOPHILIZED COMPOSITIONS OF VITAMIN D3

Abstract

The present invention relates to novel pharmaceutical compositions of Vitamin D3 containing nanoparticles, offering significant advancements in the formulation and delivery of Vitamin D3. The invention discloses a stable lyophilized formulation of Vitamin D3 within a physiological nano-particulate system, suitable for filling in unitary container closure packs. This invention addresses the challenges associated with Vitamin D3's instability to light, humidity, acids, oxidation, and heat, by incorporating polysorbate as a solubilizing agent and mannitol as a stabilizing and bulking agent. The invention is applicable to both oral and parenteral formulations, providing aqueous solutions that form nanodispersions upon reconstitution. The formulations as per the present invention are anti-microbial preservative-free, sugar-free, glycol-free, and ethanol-free, ensuring improved patient compliance and therapeutic efficacy.

Specification

Description:FIELD OF THE INVENTION:

The present invention relates to novel pharmaceutical compositions, specifically stable lyophilized formulations of Vitamin D3 containing nanoparticles, designed for both oral and parenteral administration. These formulations are developed within a physiological nano-particulate system, which enhance the solubility, stability, and bioavailability of Vitamin D3. The invention further solves challenges associated with fat-soluble vitamins, particularly the stabilization of Vitamin D3 against light, humidity, acids, oxidation, and heat, while also provide improved patient compliance through unitary container closure packs.

BACKGROUND OF THE INVENTION:

Vitamin D3, also known as cholecalciferol, is a fat-soluble vitamin crucial for the maintenance of calcium and phosphate homeostasis in the body. It plays a vital role in bone health by promoting the absorption of calcium and phosphorus from the intestines, reabsorption in the kidneys, and the regulation of parathyroid hormone levels. Deficiency in Vitamin D3 is a widespread health concern globally, affecting millions of individuals across all age groups. Inadequate levels of Vitamin D3 lead to conditions such as rickets in children, osteomalacia in adults, and contributes to osteoporosis and increased fracture risk in the elderly.

Vitamin D3 deficiency also has broader implications, including a potential increase in the risk of cardiovascular diseases, diabetes, and certain cancers. Moreover, emerging research suggests a link between low Vitamin D3 levels and autoimmune disorders, depression, and compromised immune function. Despite its critical importance, maintaining optimal Vitamin D3 levels in the body is challenging due to limited dietary sources, reduced synthesis in the skin due to inadequate sun exposure, and the instability of Vitamin D3 in various formulations.

The instability of Vitamin D3 poses a significant challenge in the formulation of pharmaceutical products. Vitamin D3 is highly sensitive to light, heat, humidity, acids, and oxidation, which lead to its degradation and loss of potency. This instability necessitates the development of pharmaceutical preparations that effectively stabilize Vitamin D3, ensuring its efficacy and shelf life. Traditional formulations, particularly those in liquid or oily forms, have limitations, including poor patient compliance, variable absorption rates, and difficulties in maintaining consistent dosage.

IN201641025758 provides to a stable nano-dispersion comprising an aqueous dispersion medium, a pharmaceutical active compound, a surfactant, a sugar-free stabilizing vehicle base, and a method of preparing the same. The invention uses stevia or alpha glucosyl stevioside as a sugar-free stabilizing agent to enhance stability and control particle size. The nanodispersion is stable for at least 6 months when stored in blow fill tubes made of polypropylene, polyethylene, or mixtures thereof.

CN103520102 discloses to a vitamin D nanoemulsion and a method for its preparation. The nanoemulsion comprises vitamin D and Tween-80 in a specific mass ratio, along with water. The preparation method involves mixing vitamin D and Tween-80 to form an oil phase, followed by the addition of water and ultrasonic treatment to obtain the nanoemulsion. The invention claims that the method is simple, efficient, and improves the water solubility and stability of vitamin D in the nanoemulsion.

IN431119 claims a stable aqueous parenteral composition comprising Vitamin D3 at a high concentration. The composition is substantially free from alcohol and oil, and exhibits enhanced bioavailability. The invention aims to overcome the limitations of prior art oily injectable formulations of Vitamin D3, which cause pain, erythema, swelling, nodule formation at the injection site, and delayed release of the active ingredient. The composition is prepared by mixing Vitamin D3, Glycofurol, a surfactant, and optionally water, a chelating agent, an antioxidant, and a buffer. The invention also provides the use of the composition for the treatment and prevention of vitamin D deficiency and insufficiency.

WO2011/013138 provides a stable, lyophilized, parenteral multivitamin formulation comprising Retinol palmitate, Cholecalciferol, Tocopherol, Ascorbic acid, Nicotinamide, Panthenol, Pyridoxine, Riboflavin, Thiamine, Folic acid, Biotin, Cyanocobalamine and Vitamin K. To address the instability of certain vitamins in aqueous solutions, the formulation incorporates Glycine, Glycocholic Acid, and Soyabean Lecithin as stabilizing agents. The invention presents a process for manufacturing this formulation, involving the preparation of an aqueous phase, a micellar system, homogenization, and lyophilization. The formulation is intended for parenteral administration and is designed to provide a stable and effective multivitamin supplement.

In response to these challenges, the present invention introduces a novel approach to stabilizing Vitamin D3 by incorporating it into a lyophilized formulation within a physiological nano-particulate system. This innovative formulation not only enhances the stability of Vitamin D3 but also improves its bioavailability and patient compliance. The nano-sized particles facilitate better absorption of Vitamin D3, ensuring rapid and consistent delivery of the active ingredient to the bloodstream. The lyophilized form further enhances the stability of Vitamin D3, protecting it from environmental factors that lead to degradation.

The pharmaceutical compositions of the present invention are designed for both oral and injectable use, offering versatility in administration based on patient needs. The oral formulation is particularly advantageous due to its aqueous nature, which upon reconstitution forms a nano-sized dispersion of Vitamin D3. This nano dispersion significantly increases the bioavailability of Vitamin D3, allowing for more efficient absorption compared to traditional formulations. Additionally, the lyophilized form of the oral formulation ensures long-term stability, making it suitable for storage and transport under various conditions.

For parenteral use, the present invention offers an aqueous-based formulation that, upon reconstitution, forms a nano dispersion of Vitamin D3. Injectable formulations typically provide a rapid rise in serum concentrations of Vitamin D3, which is particularly beneficial in clinical settings where immediate therapeutic effects are required. The lyophilized injectable formulation enhances the stability of Vitamin D3, reducing the risk of degradation during storage and handling. Moreover, the aqueous-based injectable formulation avoids the drawbacks associated with oil-based preparations, such as pain, erythema, swelling, nodule formation at the injection site, and delayed release of the active ingredient.

The present invention also includes the development of both single-dose and multiple-dose formulations, catering to the diverse needs of patients across different age groups. The single-dose formulation is particularly beneficial for pediatric and geriatric patients, where precise dosing is critical. The flexibility in dosing provided by the present invention ensures that patients receive the optimal amount of Vitamin D3 based on their specific requirements, thereby enhancing therapeutic outcomes.

One of the significant advantages of the present invention is its contribution to improved patient compliance. The oral formulation is designed to be preservative-free, sugar-free, glycol-free, and ethanol-free, addressing the concerns of patients who are sensitive to these additives. Additionally, the formulation is free from stabilizers and lactose, making it suitable for patients with lactose intolerance or those who prefer minimal excipient exposure. The pleasant taste and easy administration of the oral formulation further encourage adherence to the prescribed treatment regimen.

From an economic perspective, the present invention offers substantial benefits in terms of production and scalability. The formulation is developed in a powder form, which simplifies the manufacturing process compared to other dosage form. This streamlined production process reduces costs, making the formulation more affordable and accessible to a broader population. The use of lyophilization also extends the shelf life of the product, reducing wastage and ensuring that patients have access to potent and effective Vitamin D3 formulations.

In conclusion, the present invention represents a significant advancement in the field of Vitamin D3 pharmaceutical formulations. By addressing the challenges of stability, bioavailability, and patient compliance, the novel lyophilized Vitamin D3 formulation in a physiological nano system offers a reliable and effective solution for the prevention and treatment of Vitamin D3 deficiency. The dual application of the formulation for both oral and injectable use further enhances its versatility, making it suitable for a wide range of clinical settings. The incorporation of advanced excipients and the meticulous development process ensure that the final product meets the highest standards of quality, efficacy, and safety.

OBJECTIVE OF THE INVENTION:

The primary objective of the present invention is to develop a novel and stable pharmaceutical composition of Vitamin D3 containing nanoparticles, designed for enhanced bioavailability and stability.

The prime objective of the present invention is to create a lyophilized Vitamin D3 formulation within a physiological nano-particulate system, suitable for both oral and injectable administration, thereby addressing the instability issues related to light, humidity, acids, oxidation, and heat.

One objective of the present invention is to provide a process for producing a freeze-dried pharmaceutical formulation of Vitamin D3 that ensures the solubilization of fat-soluble compounds and their salts, leading to a stable end product.

One more objective of the present invention is to enhance the dissolution profile and therapeutic efficacy of Vitamin D3 through the use of solubilizing agents, which facilitate the formation of a stable nano dispersion upon reconstitution.

Another objective of the present invention is to achieve a pharmaceutical composition that is preservative-free for single-use applications, providing a safer and more patient-friendly option for both pediatric and geriatric populations.

Yet another objective of the present invention is to improve the ease of manufacturing and scalability by designing the composition in a lyophilized form, thereby reducing production costs and ensuring long-term stability.

A further objective of the present invention is to ensure that the pharmaceutical compositions are free from ethanol and glycol, making them suitable for patients with sensitivities or restrictions against these components.

Another objective of the present invention is to provide a parenteral formulation of Vitamin D3 that is oil-free, reducing the risk of pain, erythema, and nodule formation at the injection site while ensuring rapid serum concentration increases.

Yet another objective of the present invention is to enable the production of aqueous-based oral formulations that offer an immediate rise in serum Vitamin D3 levels, thus providing an alternative to slow-release, oil-based preparations.

One more objective of the present invention is to develop a versatile formulation that allows for the adjustment of dosage strength through dilution, catering to varying patient needs from pediatric to geriatric use, all while maintaining the integrity and efficacy of the active ingredient.

SUMMARY OF THE INVENTION:

The present invention relates to novel pharmaceutical compositions of Vitamin D3 containing nanoparticles, specifically focusing on a stable lyophilized Vitamin D3 formulation in a physiological nano-particulate system. This formulation is suitable for both oral and injectable administration, offering enhanced stability, bioavailability, and patient compliance.

The primary aspect of the present invention is to provide a stable lyophilized pharmaceutical composition of Vitamin D3 that maintains its stability against light, humidity, acids, oxidation, and heat.

One aspect of the present invention is to develop a Vitamin D3 formulation that forms a nano-dispersion upon reconstitution with water, thereby enhancing the bioavailability of Vitamin D3.

One more aspect of the present invention is to provide a lyophilized Vitamin D3 formulation that is preservative-free and suitable for single-use, thereby minimizing the risk of contamination and extending shelf life.

Another aspect of the present invention is to provide an oral pharmaceutical composition of Vitamin D3 that is aqueous-based, sugar-free, glycol-free, and ethanol-free, making it suitable for a wide range of patients, including those with specific dietary restrictions.

Yet another aspect of the present invention is to develop an injectable formulation of Vitamin D3 that is oil-free, thereby reducing the risk of pain, erythema, swelling, and nodule formation at the injection site, while also offering rapid serum concentration.

A further aspect of the present invention is to offer a process for preparing the lyophilized Vitamin D3 formulation that includes steps for dissolving the active ingredient and excipients in a controlled manner, followed by sterile filtration and lyophilization, ensuring the stability and sterility of the final product.

An additional aspect of the present invention is to provide a formulation that is filled in unitary container closure systems, ensuring precise dosing and easy administration for both oral and injectable forms.

Another aspect of the present invention is to utilize polysorbate 80 or polysorbate 20 or mixture there of as a solubilizing agent in the Vitamin D3 formulation, which enhances the solubility and stability of the active ingredient in the final product.

One more aspect of the present invention is to include antioxidants such as EDTA and BHT in the formulation to further protect the Vitamin D3 from degradation during storage and use.

Yet another aspect of the present invention is to provide a scalable manufacturing process for the Vitamin D3 formulation, allowing for efficient production while maintaining the quality and consistency of the product.

DETAILED DESCRIPTION OF THE INVENTION:

The following detailed description of the present subject matter the various embodiments. These embodiments are described in sufficient detail to enable those skilled in the art to practice the present subject matter. Other embodiments may be utilized and changes may be made without departing from the scope of the present subject matter.

References to "an", "one", or "various" embodiments in this disclosure are not necessarily to the same embodiment, and such references contemplate more than one embodiment. The following detailed description is, therefore, not to be taken in a limiting sense, and the scope is defined only by the appended claims, along with the full scope of legal equivalents to which such claims are entitled.

According to the present invention, "sugar-free" means less than 0.5 g of sugars per Reference Amount Customarily Consumed (RACC) and per labeled serving (or, for meals and main dishes, less than 0.5 g per labeled serving). The glycemic index of Mannitol is zero where as sucrose glycemic index is 100. With respect to the definition of sugar-free and considering glycemic index, the present invention is sugar-free and Mannitol is a polyol that is used as an artificial sweetener with a combination of sucralose, aspartame, sodium saccharin, or stevia or a mixture thereof.

Antimicrobial preservatives are substances added to nonsterile dosage forms to protect them from microbiological growth or from microorganisms that are introduced inadvertently during or subsequent to the manufacturing process. In the case of sterile articles packaged in multiple-dose containers, antimicrobial preservatives are added to inhibit the growth of microorganisms that may be introduced from repeatedly withdrawing individual doses.


The present invention relates to novel pharmaceutical compositions of Vitamin D3 containing nanoparticles, specifically designed to be stable in a lyophilized form within a physiological nano-particulate system. These compositions are suitable for both oral and injectable administration, providing enhanced bioavailability, stability, and ease of use. The invention addresses the instability of Vitamin D3 to light, humidity, and oxidation by creating a robust formulation that is filled into unitary container closure systems.

The primary embodiment of the present invention is a lyophilized Vitamin D3 composition that maintains its stability and efficacy over extended periods. This composition includes a solubilizing agent, a stabilizing agent, and a bulking agent, providing stability and bioavailability to the formulation of the present invention.

One embodiment of the present invention is the use of polysorbates, specifically polysorbate 80 or polysorbate 20, as solubilizing agents in the Vitamin D3 composition. These nonionic surfactants enhance the solubility of Vitamin D3, ensuring a uniform and stable solution before lyophilization. The amount of Vitamin D3 as per the present invention is in the range of 200 IU to 600000 IU.

One more embodiment of the present invention is the inclusion of mannitol as both a stabilizing and bulking agent in the formulation. Mannitol provides the necessary bulk for lyophilization while also stabilizing the Vitamin D3, preventing degradation during processing and storage. Further, mannitol also works as cryo-protectant that protect the degradation of Vitamin D3 during lyophilization process.

Another embodiment of the present invention is the method of preparing the lyophilized formulation, which involves dissolving vitamin D3 into the solubilizing agent polysorbate 80, followed by the sequential addition of the stabilizing and bulking agents. The process ensures complete dissolution of all components before sterilization and lyophilization.

A further embodiment of the present invention is the ability to adjust the pH of the formulation using a sodium hydroxide or ascorbic acid solution. This adjustment is critical for maintaining the stability and bioavailability of Vitamin D3 within the lyophilized product.

Another embodiment of the present invention is the incorporation of preservatives such as sodium methyl paraben, sodium propyl paraben, sodium benzoate, and potassium sorbate in the oral formulation. These preservatives ensure the microbiological stability of the formulation, particularly after reconstitution.

Yet another embodiment of the present invention is the creation of sugar-free and alcohol-free oral formulation. This design makes the formulation suitable for a wide range of patients, including those with dietary restrictions or sensitivities.

A further embodiment of the present invention is the development of a parenteral formulation that forms a nanodispersion upon reconstitution with water. This injectable form provides a rapid rise in serum Vitamin D3 levels, enhancing its therapeutic efficacy compared to traditional oil-based injections.

Another embodiment of the present invention is the elimination of oil from injectable formulation. This feature minimizes injection site reactions such as pain, erythema, and swelling, which are commonly associated with oil-based injections.

The pharmaceutical composition as per the present invention comprises a lyophilized Vitamin D3 formulation within a physiological nano system, wherein the composition for oral and parenteral administration.

The pharmaceutical composition as per the present invention further comprises one or more solubilizing agents selected from polysorbate 80 (Commercially available as a Tween 80), polysorbate 20, polysorbate 40, polysorbate 60, polyoxyl 40, hydrogenated castor oil, and the like. The amount of solubilizing agent according to the present invention is in the range of 2% to around 60% w/w. The % amount of amount given in the present invention in % w/w.

The pharmaceutical composition as per the present invention also comprises one or more stabilizing agent and bulking agent selected from mannitol, sorbitol and xylitol. The amount of bulking agent is 30-96%.

The pharmaceutical composition as per the present invention further comprises one or more preservatives selected from sodium methyl paraben in the amount 0.4-6%, sodium propyl paraben in the amount 0.1-1.0%, sodium benzoate in the amount 0.5-4.8%, and potassium sorbate in an amount 0.1-1.0%.

The pharmaceutical composition as per the present invention also comprises one or more pH adjusting agents selected from using sodium hydroxide or ascorbic acid.

The pharmaceutical composition as per the present invention is sugar-free and alcohol-free.
The pharmaceutical composition as per the present invention comprises the steps of dissolving Vitamin D3 into the solubilizing agent in polysorbate, adding the stabilizing and bulking agents, adjusting the pH, sterilizing the solution, and lyophilizing the mixture.

The pharmaceutical composition as per the present invention provides an injectable formulation forms a nanodispersion upon reconstitution with water.

The pharmaceutical composition as per the present invention is stored in a unitary container closure system.

The pharmaceutical composition as per the present invention is oil-free, reducing the risk of injection site reactions.

The oral formulation as per the present invention is an aqueous formulation, enhancing absorption and bioavailability. Upon reconstitution with water, it forms nano-sized particles, further increasing absorption. The lyophilized form ensures stability and prevents degradation. Higher doses are administered in a palatable form. The single dose is divided into multiple strengths by varying dilution, accommodating pediatric to geriatric needs. The formulation is preservative-free for single use, minimizing potential side effects. It is also sugar-free and glycol-free, suitable for patients with specific dietary restrictions. Additionally, it is ethanol/alcohol-free, accommodating individuals who cannot tolerate these substances.

The parenteral formulation as per the present invention is also an aqueous formulation, providing a rapid rise in serum concentration compared to oily preparations. Reconstitution with water results in the formation of nano-sized particles for increased absorption. The lyophilized form ensures stability and prevents degradation. The dose is reduced based on individual requirements and bioavailability. The formulation is antimicrobial preservative-free for single use, minimizing potential side effects. It is oil-free, avoiding pain, erythema, swelling nodule formation at the injection site, and delayed release of the active ingredient. Aqueous-based injections provide a more rapid release of the drug compared to oil-based preparations.

According to the present invention, the Vitamin D3 composition can be reconstituted with purified water, water for injection, 0.9% sodium chloride, 5% dextrose solution, or a mixture thereof. The composition of the present invention before the reconstitution is lyophilized powder or cake.

Pharmaceutically acceptable excipients are included in the composition to ensure stability, efficacy, and ease of administration. These excipients include anti-oxidants, bulking agents, surfactants, preservatives, sweeteners and pH-adjusting agents or mixtures thereof.

Sweeteners are selected one or more from sucralose, aspartame, acesulfame potassium, saccharin, neotame and stevia leaf extract or like or mixture thereof.

Anti-oxidant is selected one or more from ethylenediaminetetraacetic acid (EDTA) in the amount of 0.005-0.5%, butylated hydroxytoluene (BHT) in the amount of 0.002-0.0058%, ascorbic acid (Vitamin C), tocopherols (Vitamin E) or like or mixture thereof.

Bulking agent or cryo-protecting agent are selected one or more from mannitol, sorbitol and xylitol or like or mixture thereof.

Antimicrobial preservatives or stabilizers are selected one or more from Potassium sorbate, sodium benzoate, propyl paraben sodium, methyl paraben sodium, benzyl alcohol, chlorhexidine and benzalkonium chloride or like or mixture thereof.

pH adjusting agents are selected one or more from sodium hydroxide and ascorbic acid or like or mixture thereof.

Surfactants or solubilizing agent or solubilizer are selected one or more from Sodium lauryl sulfate, Polysorbate 80, Polysorbate 20, Sorbitan monolaurate, Sorbitan monooleate, Sorbitan monopalmitate, Tyloxapol, Sorbitan monostearate, Cetyltrimethylammonium bromide (CTAB), Poloxamer, Sodium dodecylbenzene sulfonate, Glycerol monostearate and Tween 80 or like or mixture thereof.

The present invention provides an aqueous nanoparticle composition of Vitamin D3 and solubilizing agent, wherein the composition is prepared by lyophilization process wherein (a) the composition further comprises bulking agents, antimicrobial preservatives, antioxidant, chelating agent, sweeteners, flavors and pH adjusting agent or mixtures thereof for a multi-dose unit for oral dosage form, Or (b) the composition further comprises bulking agents, antioxidant, chelating agent, sweeteners, flavors and pH adjusting agent or mixtures thereof for a single-dose unit for oral dosage form, Or (c) the composition further comprises bulking agents, antioxidant, chelating agent, with or without antimicrobial preservatives, and pH adjusting agent or mixtures thereof for the parenteral dosage form.

According to the present invention, an aqueous nanoparticle composition is oil-free, alcohol-free, and glycol-free composition.

According to the present invention, an aqueous nanoparticle composition is antimicrobial preservative-free composition for single-use dosage units.

A process for the manufacturing of an aqueous nanoparticle composition according to the present invention, wherein the composition is for oral administration, wherein the said process comprises steps of; (a) preparing vitamin D3 and antioxidant solution by dissolving in non-ionic surfactant or a mixture of surfactant (b) preparing solution by mixing with water for injection or purified water (c) optionally adding bulking agents, with or without antimicrobial preservatives, antioxidant, chelating agent, sweeteners, flavors, and pH adjusting agent to the solution step (a) (d) lyophilization of bulk solution obtains in step (c) and to obtain lyophilized cake or powder.

A process for the manufacturing of an aqueous nanoparticle composition according to the present invention, wherein the composition is for parenteral administration, wherein the said process comprises steps of (a) preparing vitamin D3 and antioxidant solution by dissolving in non-ionic surfactant or mixture of surfactant (b) preparing solution by mixing with water for injection (c) optionally adding bulking agents, with or without antimicrobial preservatives, antioxidant, chelating agent and pH adjusting agent to the solution step (a) (d) lyophilization of bulk solution obtains in step (c) and to obtain lyophilized cake or powder.

According to the present invention, when the composition is for oral administration, the flavoring agent is added prior to the lyophilization step.

The composition of the present invention for oral administration has the following advantages: aqueous composition, formation of nano form when reconstituting the powder with water, has better bioavailability, increased stability, a higher dose can be given in the palatable form, a single dose can be divided into multiple strengths i.e. from paediatric to geriatric by varying dilution, antimicrobial preservative-free for single use, sugar-free, PEG/PG-Glycol free and ethanol/alcohol-free.

The composition of the present invention for parenteral administration has the following advantages: aqueous composition, formation of nano form when reconstituted the powder with water, aqueous-based injection is also found to provide a rapid rise in serum concentration as compared to oily preparation, increased stability, rapid onset of action, antimicrobial preservative-free for single use, oil-free, alcohol/ethanol-free.

EXAMPLES

Having described the invention with reference to certain preferred embodiments, other aspects become apparent to one skilled in the art from consideration of the specification. The invention is further defined by reference to the following examples describing in detail by the preparation of the compounds of the invention. It is apparent to those skilled in the art that many modifications, both to materials and methods, are practiced without departing from the scope of the invention. The following examples are provided for illustrative purpose only and these examples are in no way limitative on the present invention.

EXAMPLE 1: SOLUBILITY STUDY OF VITAMIN D3 IN DIFFERENT SOLVENTS

Approximately 50 mg of Vitamin D3 was added to a suitable container. An initial volume of 1.0 mL of the solvent was added, and the mixture was stirred for 5 minutes. The appearance of the solution was recorded. If the Vitamin-D3 was fully dissolved, the solubility was reported as > 50 mg/mL.

If the Vitamin-D3 was not fully dissolved, then the solution was heated up to 45°C for 30 min, after heating if clear solution, the solubility is reported as >50 mg/mL.

The results are shown in Table 1 below.
Solvent Observation Estimated Solubility (in mg/mL)
Ethanol completely dissolved after 1 mL without heating > 50 mg/ml
PEG 400 No significant dissolution was observed up to 5 Ml < 50 mg/ml
Polysorbate 80 completely dissolved in 1 mL after heating at 45°C for 30 min > 50 mg/ml
Coconut oil completely dissolved in 1 mL after heating at 45°C for 30 min > 50 mg/ml
Span 80 No significant dissolved in 1 mL after heating at 45°C for 30 min < 50 mg/ml
Span 60 No significant dissolved in 1 mL after heating at 45°C for 30 min < 50 mg/ml

From the suitable solvents, as the object of the present invention is to provide an aqueous composition, Polysorbate 80 was found suitable for use in the present invention.

EXAMPLE 2: COMPOSITION AS PER PRESENT INVENTION FOR PALATABILITY TEST

The invention described herein comprises in various objects and their description as mentioned above, with respect to characteristics and processes adopted. While these aspects are emphasized in the invention, any variations of the invention described above are not to be regarded as departure from the spirit and scope of the invention as described.
Sr. No Ingredients Name Quantity per Unit for 10 unit
1 Vitamin D3 1.5 mg 15 mg
2 Tween 80 40 mg 400 mg
3 Sucralose 10 mg 100 mg
4 EDTA 0.25 mg 2.5 mg
5 Butylated hydroxytoluene 0.058 mcg 0.58 mcg
6 Mannitol Q.S to 1 gm 10 gm
7 Sodium Hydroxide Q.S to pH adjustment 5.0 to 7.0 1.0 gm
8 Ascorbic acid Q.S to pH adjustment 5.0 to 7.0 1.0 gm
9 Orange Flavour Q.S 10 mg
10 Purified Water Q.S to Q.S to 5.0 mL Q.S to 50.0 mL

Process for preparation:
1. An accurately weighed amount of Polysorbate 80 was taken and was heated up to 40°C±5°C.
2. Butylated hydroxytoluene was dissolved in the solution of step 1.
3. Vitamin D3 was added and stirred until completely solubilized.
4. Water for injection was taken and heated at a temperature of 40°C±5°C.
5. Mannitol was added slowly with continuous stirring and was dissolved.
6. EDTA calcium was added slowly with continuous stirring and dissolved.
7. Orange flavour was added slowly with continuous stirring and dissolved.
8. Make-up volume up to batch size was made and checked for palatability test.
9. The solution was filtered and lyophilized. (Lyophilization process: The freeze-drying process is monitored by thermocouple probes of representative bulk trays. trays are loaded into the freeze dryer and the temperature is gradually reduced until all thermocouples reach -40 °C. After the desired time has elapsed, the air in the chamber is evacuated and the temperature gradually increased until the shelf temperature is approximately +45 °C. The samples are held at about +45 °C for 20-28 hours. The chamber is then restored to atmospheric pressure with filtered Nitrogen and the trays are unloaded from the freeze dryer.)
The palatability test of the composition obtained in Example 2:

Evaluation of Taste of tween 80 Taste was checked by time intensity method. For this purpose, 6 human volunteers were selected. In this method, a sample equivalent to a normal dose 0.5 mg was held in the mouth for 60 seconds and volunteers were asked to evaluate the tween 80 for taste. Bitterness levels were recorded at 30 sec. The bitterness level was recorded before lyophilization and after lyophilization, using a numerical scale (3- Strong Bitter, 2 - Moderate Bitter, 1 -Slight Bitter, X - Threshold Bitter, 0 - No Bitter). These volunteers were instructed not to swallow the solution, which was placed in the mouth. They were instructed to thoroughly keep their mouth. Results are provided in the following table.

Volunteer number Parameter Before lyophilization
(Scale) After lyophilization
1 Taste evaluation 3 0
2 Taste evaluation 3 0
3 Taste evaluation 2 0
4 Taste evaluation 3 0
5 Taste evaluation 3 0
6 Taste evaluation 2 0

Surprisingly it was found that the composition obtained by the lyophilization process is using tween 80 is completely palatable.

EXAMPLES OF ORAL COMPOSITION (Example ORL1 - ORL7):

EXAMPLE ORL1:
Sr. No Ingredients Name Quantity per Unit For 30 unit
1 Vitamin D3 1.5 mg 45 mg
2 Tween 80 40 mg 1200 mg
3 Potassium sorbate 0.05 mg 1.5 mg
4 Sodium Benzoate 0.02 mg 0.6 mg
5 Sucralose 10 mg 300.0 mg
6 Orange Flavor Q.S Q.S
7 EDTA 0.25 mg 7.5 mg
8 Methyl paraben sodium 1.2 mg 36.0 mg
9 BHT 0.058 mcg 1.74 mcg
10 Propyl paraben sodium 0.12 mg 3.6 mg
11 Mannitol Q.S to 1.0 gm 30.0 gm
12 Sodium Hydroxide Q.S to pH adjustment 5.0 to 7.0 1.0 gm
12 Ascorbic acid Q.S to pH adjustment 5.0 to 7.0 1.0 gm
13 Purified Water Q.S to Q.S to 5.0 mL Q.S to 150.0 mL

Process for preparation:
1. Take accurately weighed amount of Polysorbate 80 and make it clear to heat up to 40°C±5°C.
2. Dissolve BHT in it with stir for 5 minutes till clear.
3. Add Vitamin D3 and stir until completely solubilized.
4. Take water for injection heat at a temperature of 40°C±5°C.
5. Add Mannitol slowly with continuous stirring and dissolve it.
6. Add EDTA calcium slowly with continuous stirring and dissolve it.
7. Add Sodium methyl paraben, Sodium propyl paraben, Sodium benzoate and potassium sorbate and Sweetener in it with continuous stirring and dissolve it
8. Adjust pH using 1% sodium Hydroxide solution/1% Ascorbic acid solution if required.
9. Make up volume up to batch size.
10. Filter and lyophilize the solution of step 9. (Lyophilization process: The freeze-drying process is monitored by thermocouple probes of representative bulk trays. trays are loaded into the freeze dryer and the temperature is gradually reduced until all thermocouples reach -40 °C. After the desired time has elapsed, the air in the chamber is evacuated and the temperature gradually increased until the shelf temperature is approximately +45 °C. The samples are held at about +45 °C for 20-28 hours. The chamber is then restored to atmospheric pressure with filtered Nitrogen and the trays are unloaded from freeze dryer.)
11. Collect the powder and shift it through 20# sieve.
12. Fill the dry powder in a bottle.

The samples were analyzed in the form of nanodispersion. Purified water was slowly added to the bottle containing the drug concentrate (0.02 mg drug) with gentle shaking to get a transparent nanodispersion of the drug having a dilution of 0.02 mg/ml. The nanodispersion was analyzed for the following tests: Appearance, Assay of Drug, pH (Mettler Toledo-five easy, pH Meter), and Particle size (Nano-ZS, Malvern Particle size analyzer). The analytical test results are provided below.

Sr. No. Parameter Results
1 Appearance Clear light orange colour solution
2 Assay 102.10 %
3 pH 5.99
4 Particle size 35 nm (Z-Avg)

EXAMPLE ORL2:

Sr. No Ingredients Name Quantity per Unit for 30 unit
1 Vitamin D3 1.5 mg 45 mg
2 Tween 80 40 mg 1200 mg
3 Glycofurol 5 mg 150 mg
4 Potassium sorbate 0.05 mg 1.5 mg
5 Sod. Benzoate 0.02 mg 0.6 mg
6 Sucralose 10.0 mg 300.0 mg
7 Orange Flavour Q.S Q.S
8 EDTA 0.25 mg 7.5 mg
9 Methyl paraben sodium 1.2 mg 36.0 mg
10 BHT 0.058 mcg 1.74 mcg
11 Propyl paraben sodium 0.12 mg 3.6 mg
12 Mannitol Q.S to 1 gm 30.0 gm
12 Sodium Hydroxide Q.S to pH adjustment 5.0 to 7.0 1.0 gm
13 Ascorbic acid Q.S to pH adjustment 5.0 to 7.0 1.0 gm
14 Purified Water Q.S to Q.S to 3.0 mL 90

The process for preparation is similar to example ORL1. The only difference is the addition of glycofurol with tween 80.

The analytical study results of example ORL2:

Sr. No Parameter Results
1 Appearance Clear light orange colour solution
2 Assay 99.56 %
3 pH 6.05
4 Particle size 39 nm (Z-Avg)

EXAMPLE ORL3:

Sr. No Ingredients Name Quantity per Unit for 30 unit
1 Vitamin D3 1.5 mg 45 mg
2 Tween 80 40 mg 1200 mg
3 Kolliphore Rh-40 5.0 mg 150 mg
4 Potassium sorbate 0.05 mg 1.5 mg
5 Sod. Benzoate 0.02 mg 0.6 mg
6 Sucralose 10 mg 300 mg
7 Orange Flavour Q.S Q.S
8 EDTA 0.25 mg 7.5 mg
9 Methyl paraben sodium 1.2 mg 36 mg
10 BHT 0.058 mcg 1.74 mcg
11 Propyl paraben sodium 0.12 mg 3.6 mg
12 Mannitol Q.S to 1 gm 30.0 gm
12 Sodium Hydroxide Q.S to pH adjustment 5.0 to 7.0 1.0 gm
13 Ascorbic acid Q.S to pH adjustment 5.0 to 7.0 1.0 gm
14 Purified Water Q.S to Q.S to 5.0 mL Q.S to 150.0 mL

The process for preparation is similar to example ORL1. The only difference is the addition of Kolliphore Rh-40 with tween 80.

The analytical study results of example ORL3:

Sr. No Parameter Results
1 Appearance Clear light orange colour solution
2 Assay 100.74 %
3 pH 6.04
4 Particle size 38 nm (Z-Avg)

EXAMPLE ORL4:

Sr. No Ingredients Name Quantity per Unit for 30 unit
1 Vitamin D3 1.5 mg 45 mg
2 Kolliphore Rh-40 40 mg 1200 mg
3 Potassium sorbate 0.05 mg 1.5 mg
4 Sod. Benzoate 0.02 mg 0.6 mg
5 Sucralose 10 mg 300 mg
6 Orange Flavour Q.S Q.S
7 EDTA 0.25 mg 7.5 mg
8 Methyl paraben sodium 1.2 mg 36 mg
9 BHT 0.058 mcg 1.74 mcg
10 Propyl paraben sodium 0.12 mg 3.6 mg
11 Mannitol Q.S to 1 gm 30 gm
12 Sodium Hydroxide Q.S to pH adjustment 5.0 to 7.0 1.0 gm
12 Ascorbic acid Q.S to pH adjustment 5.0 to 7.0 1.0 gm
13 Purified Water Q.S to Q.S to 5.0 mL Q.S to 150.0 mL

The process for preparation is similar to example ORL1. The only difference is the addition of Kolliphore Rh-40 instead of tween 80.

EXAMPLE ORL5: Same composition as of example ORL1, The only difference is the addition of glycofurol instead of tween 80.

The analytical study results of examples ORL4 and ORL5:

Sr. No Parameter Example ORL4 Example ORL5
1 Appearance Hazy solution formed Hazy solution formed
2 Assay * *
3 pH * *
4 Particle size * *
*not clear solution was formed, so no further analyzed.

From the above study results it can be concluded that Tween 80 is a better surfactant compared to Kolliphore Rh-40 and glycofurol.

EXAMPLE ORL6

Sr. No Ingredients Name Quantity per Unit for 100 unit
1 Vitamin D3 1.5 mg 150 mg
2 Tween 80 45.0 mg 4.0 gm
3 Potassium sorbate 0.05 mg 5.0 mg
4 Sod. Benzoate 0.02 mg 2.0 mg
5 Sucralose 10 mg 1.0 gm
6 Orange Flavour Q.S Q.S
7 EDTA 0.25 mg 25.00 mg
8 Methyl paraben sodium 1.2 mg 120.00 mg
9 BHT 0.058 mcg 5.8 mcg
10 Propyl paraben sodium 0.12 mg 12.00 mg
11 Colloidal silicon dioxide 1.00 100.0 mg
12 Mannitol Q.S to 1 gm 100.0gm
12 Sodium Hydroxide Q.S to pH adjustment 5.0 to 7.0 10.0 gm
13 Ascorbic acid Q.S to pH adjustment 5.0 to 7.0 10.0 gm
13 Purified Water Q.S to Q.S to 5.0 mL Q.S to 500.0 mL

The process for preparation is similar to example ORL1.

The composition of example ORL6 was subject to the stability study, following is the stability study results:

Stability study results for the composition of Example ORL6:

Sr. No Parameter Initial
(Before Freeze drying) Initial
(after Freeze drying) 25˚C/60% RH 40˚C/75% RH
1M 3M 6M 1M 3M 6M
1 Appearance COCS* COCS COCS COCS COCS COCS COCS COCS
2 Assay 100.35 % 99.52% 99.38% 99.02% 98.12% 99.01% 98.12% 96.56%
3 pH 5.98 6.08 6.08 6.08 6.08 6.08 6.08 6.08
4 Particle size (Z-Avg) 42 nm 38 nm 42 nm 32 nm 35 nm 36 nm 45 nm 46 nm
M= Month; * - COCS=Clear orange colour solution

The above data shows that the assay of Vitamin D3 was within 92%-110% upon storage. Thus, it can be concluded that the oral composition of Example ORL6 was found to be chemically stable when stored at room temperature (25˚C/60% relative humidity) for 6 months and at 40˚C/75% relative humidity (accelerated storage stability condition) for a prolonged period of at least 6 months.

EXAMPLE ORL7:

The composition is similar to example ORL1, except methyl paraben sodium, propyl paraben sodium, sodium benzoate, and potassium sorbate (i.e. absence of these four excipients) and was prepared by a similar process as of EXAMPLE ORAL1.

The analytical study results of example ORL7:

Sr. No Parameter Before Gamma radiation After Gamma radiation
1 Appearance Clear light orange colour solution Clear light orange colour solution*
2 Assay 100.74% 97.74%
3 pH 6.04 6.04
4 Particle size 41 nm (Z-Avg) 36 nm (Z-Avg)

The above analytical study results show that the assay of Vitamin D3 was within 92%-110% upon gamma radiation. Thus, it can be concluded that the oral composition of Example ORL7 was found to be chemically stable when applied gamma radiation.
*To protect against colour changes, amber colour bottles were used for gamma radiation purpose. Therefore, it is possible to prepare a preservative-free oral composition of vitamin D3 using the above formula.

EXAMPLES OF INJECTABLE COMPOSITION (EXAMPLE INJ1 - INJ3):

EXAMPLE INJ1:

Sr. No Ingredients Name Quantity per Unit For 50 Units
1 Vitamin D3 15 mg 0.75 gm
2 Tween 80 375 mg 18.75 gm
3 Mannitol 500 mg 25.00 gm
4 EDTA 0.25 12.5 mg
5 BHT 0.058 mcg 2.9 mcg
6 Sodium Hydroxide Q.S to pH adjustment 5.0 to 7.0 5.0 gm
7 Ascorbic acid Q.S to pH adjustment 5.0 to 7.0 5.0 gm
8 Water for injection Q.S to 4 mL 200.0 ml

Process of preparation:
1. Take an accurately weighed amount of Polysorbate 80 and make it clear to heat up to 40°C±5°C.
2. Dissolve BHT in it with stir for 5 minutes till clear.
3. Add Cholecalciferol and stir until completely solubilized.
4. Take water for injection heat at a temperature 40°C±5°C and add to the above solution.
5. Add Mannitol slowly with continuous stirring and dissolve it.
6. Add EDTA calcium slowly with continuous stirring and dissolve it.
7. Adjust pH using 1% sodium Hydroxide solution / 1% Ascorbic acid solution if required.
8. Make up volume up to batch size.
9. Filter and sterile filling carried out in an aseptic area at a fill volume 4.0 mL.
10. Lyophilize it and vial sealing carried out (Lyophilization process: The freeze-drying process is monitored by thermocouple probes of representative vials. vials are loaded into the freeze dryer and the temperature is gradually reduced until all thermocouples reach -40 °C. After the desired time has elapsed, the air in the chamber is evacuated and the temperature gradually increased until the shelf temperature is approximately +45 °C. The samples are held at about +45 °C for 30-38 hours. The chamber is then restored to atmospheric pressure with filtered Nitrogen and the vials are unloaded from the freeze dryer. Vials are removed and capped with aluminium seals).

The lyophilized powder obtained in the above example was analyzed. Water for injection was slowly added to the bottle containing the drug concentrate (15 mg drug) with gentle shaking to get a transparent nanodispersion of drug having a dilution of 7.5 mg/ml. The nanodispersion was analyzed for the following tests: Appearance, Assay of Drug, pH (Mettler Toledo-five easy, pH Meter), and Particle size (Nano- ZS, Malvern Particle size analyzer)

The analytical study results of examples INJ1:

Sr. No Parameter Results (Before Freeze drying) Results (After Freeze drying)
1 Appearance Clear colourless solution White to off white lyophilised cake
2 Assay 100.98 % 99.102 %
3 pH 5.80 5.88
4 Particle size 48 nm (Z-Avg) 48 nm (Z-Avg)

Example INJ2:

Sr. No Ingredients Name Quantity per Unit For 50 Units
1 Vitamin D3 15 mg 0.75 gm
2 Tween 80 200 mg 18.75 gm
3 Mannitol 500 mg 25.00 gm
4 EDTA 0.25 12.5 mg
5 BHT 0.058 mcg 2.9 mcg
6 Sodium Hydroxide Q.S to pH adjustment 5.0 to 7.0 5.0 gm
7 Ascorbic acid Q.S to pH adjustment 5.0 to 7.0 5.0 gm
8 Acetonitrile 1.0 mL 50.0 mL
9 Water for injection Q.S to Q.S to 4 mL Q.S to 200.0 ml

The process for preparation is similar to example INJ1.

The analytical study results of example INJ2:

Sr. No Parameter Results (Before Freeze drying) Results (After Freeze drying)
1 Appearance Clear colourless solution White to off white lyophilised cake
2 Assay 101.52 % 101.02 %
3 pH 5.92 6.01
4 Particle size 45 nm (Z-Avg) 43 nm (Z-Avg)

Example INJ3:

Sr. No Ingredients Name Quantity per Unit For 100 Units
1 Vitamin D3 7.5 mg 0.75 gm
2 Tween 80 187.5 mg 18.75 gm
3 Mannitol 250 mg 25.00 gm
4 EDTA 0.125 12.5 mg
5 BHT 0.029 mcg 2.9 mcg
6 Sodium Hydroxide Q.S to pH adjustment 5.0 to 7.0 5.0 gm
7 Ascorbic acid Q.S to pH adjustment 5.0 to 7.0 5.0 gm
8 Water for injection Q.S to 2.0 mL 200.0 ml

The process for preparation is similar to example INJ1.

The composition of example INJ3 was subject to the stability study, following is the stability study results: Stability study results for the composition of Example INJ3:

Sr. No Parameter Initial 25˚C/60% RH 40˚C/75% RH
1M 3M 6M 1M 3M 6M
1 Appearance WOFL* WOFL WOFL WOFL WOFL WOFL WOFL
2 Assay 101.50% 101.30% 101.02% 100.12% 101.70% 101.08% 99.10%
3 pH 5.95 5.95 6.02 6.03 6.03 5.99 6.10
4 Particle size (Z-Avg) 45 nm 46 nm 46 nm 36 nm 44 nm 45 nm 49 nm
M= Month; * - WOFL =white to off white lyophilised cake

The above data shows that the assay of Vitamin D3 was within 92% - 110% upon storage. Thus, it can be concluded that the injectable composition of INJ3 was found to be chemically stable when stored at room temperature (25˚C/60% relative humidity) for 6 months and at 40˚C/75% relative humidity (accelerated storage stability condition) for a prolonged period of at least 6 months.
, Claims:I claim:

1. An aqueous nanoparticle composition comprising Vitamin D3 and solubilizing agents, wherein the composition is prepared by the lyophilization process.

2. An aqueous nanoparticle composition according to claim 1, wherein the solubilizing agent is non-ionic surfactant and wherein pH of the composition is 5 to 7.

3. An aqueous nanoparticle composition according to claim 1, wherein the solubilizing agent is Polysorbate 80, Polysorbate 20, Polysorbate 40, Polysorbate 60, or a mixture thereof and the solubilizing is present in an amount of 2% to around 60%.

4. An aqueous nanoparticle composition according to claim 1, wherein the composition forms a nanodispersion upon reconstitution with purified water, water for injection, 0.9% sodium chloride, 5% dextrose solution, or a mixture thereof.

5. An aqueous nanoparticle composition according to claim 1, wherein the composition is lyophilized powder prior to the reconstitution, and the said composition is for oral or parenteral administration.

6. An aqueous nanoparticle composition according to claim 1,
(a) wherein the composition further comprises bulking agents, antimicrobial preservatives, antioxidant, chelating agent, sweeteners, flavours and pH adjusting agent or mixtures thereof for a multi-dose unit for oral dosage form, Or
(b) wherein the composition further comprises bulking agents, antioxidant, chelating agent, sweeteners, flavours and pH adjusting agent or mixtures thereof for single-dose unit for oral dosage form, Or
(c) wherein the composition further comprises bulking agents, antioxidant, chelating agent, with or without antimicrobial preservatives, and pH adjusting agent or mixtures thereof for parenteral dosage form.

7. An aqueous nanoparticle composition according to claim 1,
(a) wherein the composition is oil-free or alcohol-free composition or glycol-free composition, Or
(b) wherein the composition is an antimicrobial preservative-free composition for single-use dosage unit.

8. A process for the manufacturing of an aqueous nanoparticle composition according to claim 1, wherein the composition is for oral administration, wherein the said process comprises steps of;
(a) preparing vitamin D3 and antioxidant solution by dissolving in non-ionic surfactant or mixture of surfactant
(b) preparing solution by mixing with water for injection or purified water
(c) optionally adding bulking agents, with or without antimicrobial preservatives, antioxidant, chelating agent, sweeteners, flavours and pH adjusting agent to the solution step (a)
(d) lyophilization of bulk solution of step (c) to obtain lyophilized cake or powder.

9. A process for the manufacturing of an aqueous nanoparticle composition according to claim 1, wherein the composition is for parenteral administration, wherein the said process comprises steps of;
(a) preparing vitamin D3 and antioxidant solution by dissolving in non-ionic surfactant or a mixture of surfactant
(b) preparing solution by mixing the solution of step (a) with water for injection
(c) optionally adding bulking agents, with or without antimicrobial preservatives, antioxidant, chelating agent and pH adjusting agent to the solution step (a)
(d) lyophilization of bulk solution of step (c) to obtain lyophilized cake or powder.

10. An aqueous nanoparticle composition according to claim 1, wherein the composition is for oral administration and wherein the flavoring agent is added prior to the lyophilization step.

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