Description:1
FORM-2
THE PATENTS ACT, 1970
(39 of 1970)
&
THE PATENTS RULES, 2003
COMPLETE
SPECIFICATION
(See section 10 and rule 13)
Topical Nanoemulgel Formulation of Tofacitinib Citrate and Diacerein
G D Goenka University, an Indian university of Sohna Gurugram Road,
Sohna, Haryana, India, 122103
THE FOLLOWING SPECIFICATION PARTICULARLY DESCRIBES THE INVENTION AND THE MANNER IN WHICH IT IS TO BE PERFORMED
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FIELD OF INVENTION:
The present invention relates to pharmaceutical science field, which aims at design, development and evaluation of novel topical formulation of tofacitinib citrate and diacerein.
BACK GROUND:
Arthritis is a persistent systemic auto-immune syndrome caused through redundant mobilization of inflammatory cells, i.e. monocytes, macrophages, B & T lymphocytes, and dendritic cells of synovial fluid, bringing about abnormal increment in synovial cells, novel formation of the vascular system along with cartilage destruction. Cartilage destruction is followed by distortion and damage to joints, incredibly trivial joints of appendages; other joints, including additional body parts such as eyes, lungs skin, and heart, also get affected. The typical indicators of arthritis are joint distress with inflammation and rigidity intensifying at dawn or afterwards latent phase comprising joints of similar sides (for example, knee joint or wrist joint). Rheumatoid arthritis, Osteoarthritis, and Psoriatic arthritis are widespread kinds of arthritis. Osteoarthritis is the prevailing kind of arthritis globally, affecting aged persons. Usually, it affects the joints of the knees, hands, spine, and hips by eroding the shielding cartilage of bones. Rheumatoid arthritis instigates pain, inflammation and stiffness of joints and bones and results in bone devastation. Psoriatic arthritis is a complex inflammatory illness mainly involving the musculoskeletal system and skin. Initially, psoriasis in the skin, i.e. red patches, develops later on pain and stiffness of joints occurs. The participation of many cytokines causes rheumatoid arthritis, which is an autoimmune disorder. The mechanism behind the progression of this disease is the cytokines that are essential for immunoregulation and body defence. Rheumatoid arthritis is predominantly indicated through three kinase lines, i.e. Syk (Spleen Tyrosine Kinase), JAK (Janus Kinase), in addition to p38 (Mitogen-Activated Protein Kinase).
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TNF-a is predominantly produced by macrophages and further by natural killer cells, neutrophils, and activated lymphocytes, which aid in the development of connective tissue growth factors through rapid induction of synovial fibroblasts, which incites the atypical osteoclasts causing joint impairment. TNF-a promotes the Pannus development by promoting leucocytes cascade over the synovial fluid. Both IL-1 & TNF-a are liable to disseminate the inflammatory chute, while p38 (a & ß) slows down the generation of such cytokines.
Tofacitinib, the pioneer JAKinib detected in November 2012 for RA; oral tofacitinib inhibits enormous signalling pathways of cytokines. It effectively hinders JAK 1&3, causing blockage of interleukins (IL-2, 4, 7, 9,15 & 21) and IL- 6, 10 & 11, interferons (IFN- a/ß, IFN- ?), respectively, whereas slightly inhibits JAK 2 and Tyk2. For the safe and efficacious treatment of RA, it is given twice per day in doses of 5 and 10 mg, both in monotherapy as well as a combined therapy. Tofacitinib is the paramount Jakinib being commended for RA therapy. The assets of tofacitinib are accompanied by some harmful severe upshots and vary to other biological DMARDs in possessing oral tofacitinib was found efficacious for treating RA in patients of Latin America (LA) with regulated protection lines. In Phase IIb and III clinical trials, tofacitinib was found to be efficacious at a dose of 10 mg. Using both doses, i.e. 5 & 10 mg b.i.d. significant rise in ACR (20, 50 &70) was noted in contrast to both control and methotrexate.
Diacerein (DCN) is a chondroprotective diacetyl variant of rhein that is being used to alleviate osteoarthritis. It’s a BCS class II medication with special traits like anti-oxidant as well as anti-apoptotic action. In vitro, DCN and rhein reduce TNF-a and IL-12 generation while decreasing IL-1 production in humanlike monocytes. It also suppresses the synthesis release of IL-1, reducing IL-1 related activities in experimental osteoarthritis models. DCN controls metalloprotease-13, plasminogen activator, vitamin-D3 stimulated osteocalcin, COX-2, PEG-2 in subchondral osteoblasts. In RA patients who have not responded well to methotrexate, DCN can help relieve pain, restore joint function, and also provide respite.
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Nanoemulsion evinces a high prospect in the drug delivery system due to its transparency and isotropous nature of the two phases, which are being steadied by stabilizing or surface-active agents. These also exhibit a high solubility profile of the drug compared to the conventional micelle system. NEs also have advanced absorption through the skin, extended stability and shelf life. But instead of numerous pluses, these are inadequate for the topical route because of their low viscosity and spreading coefficient. Nanoemulgel (NEG) is precisely the nanoemulsions assimilated into hydrogels ground possessing improved skin penetration. Nanoemulgels behave like a drug reservoir, affecting drug delivery to the skin based on polymer configuration and cross-linking intensity. It increases the stability of nanoemulsions by dispersing them in gel, enhancing adhesion ability, spreadability and concentration gradient. It aids in the half-life of drugs by bestowing controlled release. Nanoemulgels are patient compliant, non-toxic and non-inflammatory.
Kathuria et al. 2020, formulated proposomes of tofacitinib for transdermal delivery. The proposome was characterized for its stability, vesicle morphology, polydispersity index (PDI), zeta potential and entrapment efficiency. In addition, the skin absorption of TC through proposomes was also assessed. The proposome composition was found to affect the absorption of TC into the different skin layers.
Gorantla et al. 2023, developed hyaluronic acid-coated proglycosomes of tofacitinib. he therapeutic efficacy of TF can be improved through the high affinity of natural ligands (hyaluronic acid and chondroitin sulphate) to CD44 receptors on the macrophages or other immune cells in the dermal region. Thus, the present research work was inspired by the possibility to develop and evaluate the potential of hyaluronic acid-coated proglycosomes (HA-TF-PG) as the carrier for site-specific dermal delivery. The normal-PG (N-TF-PG) and HA-TF-PG showed particle sizes of <250 nm. The HA-TF-PG demonstrated 3.15-fold higher retention of TF in the viable dermis layers than the conventional formulation. The in vivo pharmacodynamic study, cytokines, and radiographic study on Complete
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Freud's Adjuvant-induced arthritic rat model revealed that HA-TF-PG exhibited a significant (P < 0.001) reduction in inflammation in arthritic rat's paw compared to the conventional TF. The developed HA-TF-PG treated groups showed significantly lowered CD44 levels compared to FD-gel and N-TF-PG i.e. 2.28 and 1.32-fold respectively (p < 0.001). In conclusion, The HA-TF-PG can be developed as an effective carrier for the site-specific dermal drug delivery system of TF to treat RA.
Attempt has been made by some researcher for topical delivery of tofacitinib for the treatment of rheumatoid arthritis. But oral formulations of Tofacitinib are effective but pour serious unavoidable side effects. Also, topical formulations of Tofacitinib for rheumatoid arthritis are not available in the market. Hence, formulation development and evaluation of Nanoemulgel of Tofacitinib citrate and Diacerein is done with following objectives:
OBJECTIVE OF THE INVENTION:
1. It is an object of the invention to provide a method for preparing Cilostazol Solid Dispersion Nanoparticles (SND) with Thymoquinone (TLS) (80 mg) and Curcumin tag.
2. It is another object of the invention to provide pharmaceutical composition comprising Cilostazol Solid Dispersion Nanoparticles (SND) with Curcumin and Thymoquinone (TLS) tag.
3. It is another object of the invention to method for enhancing the therapeutic effectiveness of Cilostazol, comprising preparing Cilostazol Solid Dispersion Nanoparticles (SND) with Thymoquinone (TLS) and Curcumin tag using the emulsion solvent evaporation process.
4. It is another object of the invention to develop composition provides enhanced drug delivery and controlled release properties, allowing for synergistic effects of Thymoquinone and Curcumin in various therapeutic applications.
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SUMMARY
JAK inhibitors are the new diseasemodifying agents that are highly effective in relieving the joint’s pain, stiffness and swelling. Tofacitinib citrate in oral showed good bioavailability but the adverse effects led to the idea of topical formulation of tofacitinib citrate. The physicochemical characteristics of tofacitinib citrate are also suitable for making topical formulation, for example, low molecular weight, small t1/2, and log P value. Similarly, Diacerein a medicinal agent obtained from rheinalkaloids, is effective in the management of arthritis. In order to avoid the adverse effects and gain high efficacy, the topical formulation of diacerein is developed. Oleic acid, tween 80, and propylene glycol were selected as oil, surfactant, and co-surfactant, respectively on the basis of various screening tests. Ultrasonication is a high-energy method of producing nanoemulsionswith reduced particle size. It is a method that provides nanoemulsions with small size in a short time. BBD was employed as it provides maximum effectiveness of responses related to critical factors with fewer runs. Particle size and % LE were selected as the response factors. The concentration of oleic acid, tween 80, and propylene glycol had a significant effect on both droplet size and % LE. In order to increase the stability and ease in application, the nanoemulsion formulation was dispersed in 1 % Carbopol gel. DSC of the formulations confirmed no new peak formation and amorphous nature of the drug too. The in vitro release studies exhibited a good % cumulative release of NEG as compared to NE and plain gel for both. Good rheological behavior with enhanced skin permeability was also observed for TFB-NEG and DC-NEG. The stability studies conducted at various temperatures 4 °C, 25 °C, 40 °C, indicated no significant change in the pH, viscosity, and drug content of the formulations. The anti-arthritic studies performed on the rats exhibited a significant decrease in the paw volume and diameter of the rats induced with CFA. Overall, TFB-NEG and DC-NEG formulations could be considered an alternative delivery approach for enhanced skin delivery and reduced adverse events as observed with oral formulations.
BRIEF DESCRIPTION OF FIGURES
These and other features, aspects, and advantages of the present invention will become
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better understood when the following detailed description is read with reference to the accompanying figures in which like characters represent like parts throughout the figures, wherein:
Figure 1, illustrates a view of Skin Permeation of TFB for the present invention.
Fig. 2: illustrates a view of Skin Permeation study of DCN for the present invention.
Fig.3: illustrates a view of TEM photomicrograph of DCN-NE for the present invention.
Fig.4: illustrates a view of % Cumulative Release of DCN from the plain gel, DCN-NE, and DCN-NEG for the present invention.
Fig: 5: illustrates a view of Graphical Representation of Stability Analysis of Optimized Formulation for the present invention.
Fig: 6: illustrates a view of Impacts of TFB-NEG and DCN-NEG on Paw Edema in Comparison to Standard (Nimulid gel) for the present invention.
Further, skilled artisans will appreciate that elements in the figures are illustrated for simplicity and may not have been necessarily been drawn to scale. For example, the flowcharts illustrate the method in terms of the most prominent steps involved to help to improve understanding of aspects of the present invention. Furthermore, in terms of the construction of the device, one or more components of the device may have been represented in the figures by conventional symbols, and the figures may show only those specific details that are pertinent to understanding the embodiments of the present invention so as not to obscure the figures with details that will be readily apparent to those of ordinary skill in the art having benefit of the description herein.
DETAILED DESCRIPTION:
For the purpose of promoting an understanding of the principles of the invention, reference will now be made to the embodiment illustrated in the figures and specific language will be used to describe the same. It will nevertheless be understood that no limitation of the scope of the invention is thereby intended, such alterations and further modifications in the illustrated system, and such further applications of the
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principles of the invention as illustrated therein being contemplated as would normally occur to one skilled in the art to which the invention relates.
It will be understood by those skilled in the art that the foregoing general description and the following detailed description are exemplary and explanatory of the invention and are not intended to be restrictive thereof.
Reference throughout this specification to “an aspect”, “another aspect” or similar language means that a particular feature, structure, or characteristic described in connection with the embodiment is included in at least one embodiment of the present invention. Thus, appearances of the phrase “in an embodiment”, “in another embodiment” and similar language throughout this specification may, but do not necessarily, all refer to the same embodiment.
The terms "comprises", "comprising", or any other variations thereof, are intended to cover a non-exclusive inclusion, such that a process or method that comprises a list of steps does not include only those steps but may include other steps not expressly listed or inherent to such process or method. Similarly, one or more devices or systems or elements or structures or components proceeded by "comprises... a" does not, without more constraints, preclude the existence of other devices or other systems or other elements or other structures or other components or additional devices or additional systems or additional elements or additional structures or additional components.
Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. The system, methods, and examples provided herein are illustrative only and not intended to be limiting.
The terms “a” and “an” herein do not denote a limitation of quantity, but rather denote the presence of at least one of the referenced items.
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The terms “having”, “comprising”, “including”, and variations thereof signify the presence of a component.
Now the present invention will be described below in detail with reference to the following embodiment.
Example 1
Formulation Development and Evaluation of Nanoemulgel of TFB
TFB-NEG was formulated in following two phases:
Phase 1: Preparation of TFB nanoemulsion (TFB-NE)
The NE of TFB was prepared by using high energy ultrasonication method. This method produces cavitation resulting in decreased particle size and uniform particle size distribution. Depending on the solubility analysis the oil, surfactant and cosurfactanat were used for nanoemulsion preparation as mentioned in table 1. The oil phase was prepared using 0.2% TFB and the required amount of tween 80 and propylene glycol. The aqueous phase was employed with required amount of deionized water. The oil phase was mixed with aqueous phase to make nanoemulsion using a probe ultrasonicator(ChromTech Bio-Age) for 30 minutes while maintaining outside temperature using ice jacketed bath. In, this way homogeneous nanoemulsion was formed without overheating. The prepared NEs were evaluated for particle size, %EE and %LE.
Phase 2: Preparation of TFB-NEG
NEGs containing the optimized NE of TFB were prepared using carbopol-934, triethanolamine and deionized water. The required quantities of carbopol 934 for 0.5%, 1% , and 1.5% w/v were taken and dispersed in water for 24 hr for complete and uniform swelling. Triethanolamine was added to neutralize the pH to around 5.5. Afterward, the optimized NE of TFB was mixed with the hydrogel to make it TFB loaded NEG. The
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prepared NEG formulations were evaluated for pH, viscosity, spreadability, drug content, and in-vitro drug release studies.
The TFB-NE was formulated according to BBD, and 17 formulations were made and characterized to optimize the formulation. The range of droplet size and % loading efficiency were 106.3 ± 2.8 to 247.2 ± 0.28 nm and 1.3 ± 0.97 to 18.9 ± 0.86 %, respectively. The quadratic equation for both droplet size and percent LE showed that oil content had a positive influence on droplet size and a negative influence on percent LE, whereas an increase in the extent of surfactant and co-surfactant had a negative influence on droplet size and a positive influence on percent LE. The R2 value obtained for droplet size and % LE were 0.9986 and 0.9825, respectively. The p-value for both was found to be < 0.0001, illustrating the significance of the design. The predicted and experimental values obtained for droplet size and % LE were found close. The optimized formulation was selected through BBD outcomes. The TEM analysis of the optimized formulation confirmed the nano-size range of the formulation. The optimized TFB-NE was then dispersed in 1 % carbopol 934 P gel to obtain Tofacitinib Nanoemulgel (TFB-NEG). The pH, spreadability of TFB-NEG were found to be 5.8 ± 0.2 and 6.4 ± 0.46 g cm/s. The drug content of the top, middle, and bottom layers were found to be 92.64 ± 1.28, 92.79 ± 1.83, 94.37 ± 2.76 respectively, showing that the loss of drug upon addition to NEG was insignificant. The % cumulative drug release after 24 hours was found to be 89.64 ± 0.97 % for TFB-NEG which was higher than TFB-NE and TFB-Plain gel. The Kinetics model suggested that the TFB-NEG followed the Higuchi model and non-Fickian release pattern. The Flux (µg/cm2/min), Permittivity (cm/min), and Cumulative amount per cm2 at 24 hour were 0.249 ± 0.019, 0.062 ± 0.005, and 5.25 ± 0.12 respectively.. The stability studies conducted at various temperatures 4 °C, 25 °C, 40 °C, indicated no significant change in the pH, viscosity, and drug content of the formulations. The anti-arthritic studies performed on the rats exhibited a significant decrease in the paw volume and diameter of the rats induced with CFA.
Example 2
Ex-vivo Permeation Studies
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TFB-NEG, TFB-NE, and plain TFB-gel, TFB permeability was assessed and compared (Fig.1). Table 1 enlists the permeation parameters such as flux, permittivity, cumulative amount/ cm2 after 24 hours, and the p-value. After dilution, all samples were collected at predefined intervals and analyzed using a UV spectrophotometer at 286 nm. Because of the smaller size of particles of the formulation, advanced drug absorption is possible, resulting in high bio-distribution in the affected area and a higher concentration gradient, which is necessary for successful dermal delivery—the size of the nano-formulation influences the transdermal transit. Size plays a vital role in transportation through the skin. Nanoemulsions with small particle sizes can diffuse into the epidermis and enter deep-lying skin layers; however, the larger particle size limits drug diffusion to the epidermis and internal skin layers.
Table 1: Parameters of Skin Permeation of TFB with ANOVA analysis
Formulation
Flux (µg/cm2/min)
Permittivity (cm/min)
Cumulative Amount per cm2 at 24 hour
p-value
TFB-NEG
0.249 ± 0.019
0.062 ± 0.005
5.25 ± 0.12
NEG vs. NE < 0.01; NEG vs. plain gel < 0.001
TFB-NE
0.172 ± 0.005
0.043 ± 0.002
3.87 ± 0.10
NE vs. plain gel < 0.001
Plain TFB gel
0.044 ± 0.013
0.011 ± 0.003
1.08 ± 0.02
Overall p value < 0.0001; F value 173.89
Example 3
Formulation Development and Evaluation of Nanoemulgel of DCN
The nanoemulsion of DCN (0.5 %) was prepared by using a high energy method, i.e. ultrasonication method. Depending on the solubility analysis, oleic acid, tween 80 and propylene glycol were used for nanoemulsion preparation. The placebo nanoemulsion was prepared with oil: surfactant: cosurfactant depending on the literature survey and trials. For placebo
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nanoemulsion, the oil and aqueous phases were mixed using ultrasonication for 30 minutes. Similarly, a drug-loaded nanoemulsion was prepared by dissolving the drug in the oil phase for 10 min and mixing it with the oil phase using ultrasonication. The formulations were optimized using Box-Behnken design. It provides a lesser design point compared to the central composite design; hence, it is less costly to run the design with similar factors. Unlike CCD, Box-Behnken design does not possess axial points that indicate design points within the safe operating region. According to Box-Behnken 32 design, 17 formulations were prepared with oil, surfactant, cosurfactant independent variables, droplet size, and entrapment efficiency as dependent variables. The nanoemulgel (NEG) of DCN was prepared by using carbopol 934 as a gelling agent. Carbopol can produce gel in concentrations 0.5 %, 1 %, and 1.5 %, but in 1 % concentration, the gel produces suitable for topical application and possesses good rheological assets. An accurately weighed amount of carbopol 934 was first dissolved in distilled water and then kept aside the whole night for uniform swelling. Triethanolamine in 0.3 % concentration was added for neutralization and formation of the gel matrix. Ultimately, the optimized nanoemulsion was dispersed homogeneously into the hydrogel to get DCN loaded nanoemulgel (DCN-NEG).
Example 3
Ex-vivo Permeation Studies
DCN permeability was assessed and compared for DCN-NEG, DCN-NE, and plain DCN-gel. Table 2 lists the permeation parameters such as flux, permittivity, and cumulative amount/ cm2 after 24 hours. After dilution, all samples were collected at predefined intervals, analyzed using UV spectrophotometer at 258.5 nm. Because of smaller size of particles of the formulation, advanced drug absorption is possible, which may result in high bio-distribution in the affected area and a higher concentration gradient, which is necessary for successful dermal delivery. The size of the nano-formulation influences the transdermal transit. Size plays a vital role in transportation through skin. Nanoemulsions with low particle size can diffuse into the epidermis and enter into deep-lying skin layers; however, the larger particle size limits drug diffusion to epidermis and internal skin layers.
Table 2. Parameters of Skin Permeation study of DCN with ANOVA Analysis
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Formulation
Flux (µg/cm2/min)
Permittivity (cm/min)
Cumulative Amount per cm2 at 24 hour
p-value
DCN-NEG
0.574 ±
0.574 ± 0.0160.016
0.057 ±
0.057 ± 0.0130.013
12.73 ± 0.23
12.73 ± 0.23
NEG vs NE< 0.001 ; NEG vs. plain gel < 0.001
DCN-NE
0.350 ±
0.350 ± 0.0090.009
0.035 ±
0.035 ± 0.0150.015
8.68 ± 0.18
8.68 ± 0.18
NE vs. plain gel < 0.001
Plain DCN gel
0.075 ±
0.075 ± 0.0120.012
0.008 ±
0.008 ± 0.0030.003
2.71 ± 0.01
2.71 ± 0.01
Overall p value < 0.0001
The DC-NE was formulated according to BBD, and 17 formulations were made and characterized to select the optimized formulation. The range of droplet size and % loading efficiency was 95.7 ± 2.4 to 214.8 ± 0. 31 nm and 3.1 ± 0.77 to 18.5 ± 0.21 %, respectively.
The R2 value obtained for droplet size and % LE were 0.9892 and 0.9940, respectively. The p-value for both was found to be < 0.0001, illustrating the significance of the design. The predicted and experimental values obtained for droplet size and % LE were found close. The optimized formulation was selected through BBD outcomes.
The TEM analysis of the optimized formulation confirmed the nano-size range of the formulation.
The optimized DC-NE was then dispersed in 1 % carbopol 934 P gel to obtain Diacerein Nanoemulgel (DC-NEG).
The pH, Spreadability of DC-NEG were found to be 6.1 ± 0.3 and 5.8 ± 0.7 g cm/s. The drug content of the top, middle, and bottom layers were found to be 92.79 ± 3.09, 92.93 ± 1.51, 93.54 ± 3.15, respectively and showing that the loss of drug upon addition to NEG was insignificant.
The % cumulative drug release after 24 hour was found to be 93.61 ± 2.57 % for DC-NEG which was higher than DC-NE and DC-Plain gel. The Kinetics model suggested that the DC-NEG followed the Higuchi model and non-Fickian release pattern.
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The Flux (µg/cm2/min), Permittivity (cm/min), and Cumulative amount per cm2 at 24 hour were 0.574 ± 0.016, 0.057 ± 0.013, and 12.73 ± 0.23, respectively.
The animal studies carried on Wistar rats weighing exhibited effective improvement in the paw volume and paw diameter upon application of both TFB-NEG and DC-NEG compared to Nimulid (Control) and Diseased. The p value obtained for both the formulations at 7th day was found to be < 0.001, indicating the significance of the novel topical formulations of TFB and DC for arthritis treatment.
The stability studies carried under 4 °C ± 3 °C/60% ± 5%, 25 °C ± 2 °C/60% ± 5% and 40 °C ± 2 °C /75% ± 5% RH indicated the stability of TFB-NEG and DC-NEG at 4 °C ± 3 °C/60% ± 5% and 25 °C ± 2 °C/60% ± 5% with a p value > 0.05. while the p value for 4 °C vs 25 °C was < 0.05
Example 4
Anti-inflammatory studies for TFB-NEG and DCN-NEG
The anti-inflammatory activities of TFB-NEG based on the observation of paw volume (Table 3) and paw diameter (Table 4). Figure 6 describes the pictures of rats placed in various groups.
Table 3: Mean Paw Volume vs. Time Period
Time Period (Days)
Normal (Control)
Disease
Standard (Nimulid gel)
TFB-NEG
DCN-NEG
Overall p value
F value
0
1.017 ±
0.097
1.020 ±
0.120
1.000 ±
0.07
1.064 ±
0.065
1.088 ±
0.085
0.4269ns
0.9984
1
1.023 ±
0.081
1.543 ±
0.082
1.420 ±
0.064
1.447 ±
0.056
1.453 ±
0.068
< 0.0001ns*
41.503
3
1.018±
0.103
1.624 ±
0.089
1.353 ±
0.006
1.427 ±
0.034
1.410 ±
0.054
< 0.0001 adef
54.183
5
1.015 ±
0.118
1.732 ±
0.041
1.302 ±
0.025
1.405 ±
0.026
1.388 ±
0.042
< 0.0001adef
102.55
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7
1.018 ±
0.110
1.912 ±
0.051
1.292 ±
0.022
1.365 ±
0.082
1.257 ±
0.042
< 0.0001adef
67.191
ap< 0.001,bp < 0.01, cp< 0.05, ns: non-significant, d: Disease vs standard, e: Disease vs TFB-NEG, f: Disease vs DCN-NEG, ns*: p-value is > 0.05 for Disease vs standard, Disease vs. TFB-NEG, Disease vs DCN-NEG. One-way analysis of variance (ANOVA) followed by Tukey’s Multiple Range Test were used to analyze the data statistically. The results were Mean ±SD for all six groups; also the experiments were conducted three times.
Table 4: Mean Paw diameter vs. Time Period
Time Period (Days)
Normal (Control)
Disease
Standard (Nimulid gel)
TFB-NEG
DCN-NEG
Overall p-value
F value 0 0.6717 ± 0.0325 0.655 ± 0.051 0.668 ± 0.017 0654 ± 0.025 0.662 ± 0.015 0.8040 0.4039
1
0.675 ±
0.033
0.713 ±
0.023
0.700 ±
0.028
0.734 ±
0.041
0.758 ±
0.052
0.0078 ns
4.413 3 0.675± 0.029 0.732 ± 0.025 0.688 ± 0.032 0.755 ± 0.034 0.740 ± 0.046 0.0013ns 6.195
5
0.682 ±
0.026
0.755 ±
0.026
0.687 ±
0.028
0.690 ±
0.024
0.687 ±
0.044
0.0013bdef
6.196 7 0.675 ± 0.024 0.757 ± 0.027 0.682 ± 0.028 0.682 ± 0.051 0.688 ± 0.029 0.0015bdef 6.074
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ap< 0.001, bp< 0.01, cp< 0.05, ns: non-significant, d: Disease vs standard, e: Disease vs TFB-NEG, f: Disease vs DCN-NEG. One-way analysis of variance (ANOVA) followed by Tukey’s Multiple Range Test were used to analyze the data statistically. The results were Mean ±SD for all six groups; also the experiments were conducted three times.
The paw volume and paw diameter of the CFA-induced rats were measured and compared for the corresponding treatmentgroups at intervals i.e. 0 day, 1st day, 3rd day, 5th day, and 7th day. From day 1 to day 7, a decrease in paw volume and diameter was seen for the treatment groups i.e. standard group, TFB-NEG treated group and DCN-NEG treated group. Maximum decrease in the value of paw volume was observed for the group treated with DCN-NEG (1.257 ± 0.042 ml) in contrast to TFB-NEG (1.365 ± 0.082 ml) treated group. While the reduction in paw diameter was found to be more with TFB-NEG (0.682 ± 0.051 cm) treated group as compared to DCN-NEG (0.688 ± 0.029 cm). The reduction in paw volume for DCN-NEG group suggests that the diacerein exhibited more topical penetration.
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We claim,
1. A method for preparing Topical Nanoemulgel Formulation of Tofacitinib Citrate (TFB) and Diacerein (DCN) comprising the steps of:
I. Preparation of TFB nanoemulsion (TFB-NE) and DCN-NE using high-energy ultrasonication
a) Preparing the oil phase using 0.2% TFB, 0.5% DCN, tween 80, and propylene glycol as per the solubility analysis in Table 1.
b) Preparing the aqueous phase using the required amount of deionized water.
c) Mixing the oil phase with the aqueous phase to form the nanoemulsion using a probe ultrasonicator for 30 minutes, while maintaining the outside temperature using an ice jacketed bath.
d) Forming a homogeneous nanoemulsion without overheating due to the cavitation effect resulting from high-energy ultrasonication.
II. Preparation of TFB-NEG, DCN-NEG using Carbopol-934 and triethanolamine:
a) Dispersing the required quantities of Carbopol-934 at concentrations of 0.5%, 1%, and 1.5% w/v in deionized water for 24 hours to achieve complete and uniform swelling.
b) Adding triethanolamine to the dispersed Carbopol-934 to neutralize the pH to approximately 5.5.
c) Mixing the optimized TFB and DCN nanoemulsion (NE) with the hydrogel to obtain the TFB loaded NEG formulation and DCN loaded NEG formulation.
d) Evaluating the prepared NEG formulations for pH, viscosity, spreadability, drug content, and in-vitro drug release studies.
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2. A method for optimizing the formulation of TFB (Trifluperidol) and DCN for topical application, comprising preparing a TFB and DCN loaded nanoemulsion gel (NEG) using Carbopol-934 and triethanolamine as claimed in claim 1.
3. The method for preparing Topical Nanoemulgel Formulation of Tofacitinib Citrate (TFB) and Diacerein (DCN) as claimed in claim 1, wherein Oleic acid, tween 80, and propylene glycol were selected as oil, surfactant, and co-surfactant, respectively.
4. The Topical Nanoemulgel Formulation of Tofacitinib Citrate and DiacereinMaximum prepared by a method claimed in claim 1, exhibits decrease in the value of paw volume of albino rats treated with DCN-NEG (1.257 ± 0.042 ml) in contrast to TFB-NEG (1.365 ± 0.082 ml) treated group.
5. The Topical Nanoemulgel Formulation of Tofacitinib Citrate and DiacereinMaximum prepared by a method claimed in claim 1, exhibits reduction in paw diameter of albino rats be more with TFB-NEG (0.682 ± 0.051 cm) treated group as compared to DCN-NEG (0.688 ± 0.029 cm).
Dated this 07/08/2023 G D Goenka University, Sohna Gurugram Road, Sohna, Haryana, India, 122103
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ABSTRACT
Topical Nanoemulgel Formulation of Tofacitinib Citrate and Diacerein
The present invention relates to TFB-NEG and DC-NEG formulations as an alternative delivery approach for rheumatoid arthritis with enhanced skin delivery and reduced adverse events as observed with oral formulations. The Topical Nanoemulgel Formulation of Tofacitinib Citrate and DiacereinMaximum exhibits decrease in the value of paw volume of albino rats treated with DCN-NEG (1.257 ± 0.042 ml) in contrast to TFB-NEG (1.365 ± 0.082 ml) treated group and reduction in paw diameter of albino rats be more with TFB-NEG (0.682 ± 0.051 cm) treated group as compared to DCN-NEG (0.688 ± 0.029 cm). The reduction in paw volume for DCN-NEG group suggests that the diacerein exhibited more topical penetration. , Claims:We claim,
1. A method for preparing Topical Nanoemulgel Formulation of Tofacitinib Citrate (TFB) and Diacerein (DCN) comprising the steps of:
I. Preparation of TFB nanoemulsion (TFB-NE) and DCN-NE using high-energy ultrasonication
a) Preparing the oil phase using 0.2% TFB, 0.5% DCN, tween 80, and propylene glycol as per the solubility analysis in Table 1.
b) Preparing the aqueous phase using the required amount of deionized water.
c) Mixing the oil phase with the aqueous phase to form the nanoemulsion using a probe ultrasonicator for 30 minutes, while maintaining the outside temperature using an ice jacketed bath.
d) Forming a homogeneous nanoemulsion without overheating due to the cavitation effect resulting from high-energy ultrasonication.
II. Preparation of TFB-NEG, DCN-NEG using Carbopol-934 and triethanolamine:
a) Dispersing the required quantities of Carbopol-934 at concentrations of 0.5%, 1%, and 1.5% w/v in deionized water for 24 hours to achieve complete and uniform swelling.
b) Adding triethanolamine to the dispersed Carbopol-934 to neutralize the pH to approximately 5.5.
c) Mixing the optimized TFB and DCN nanoemulsion (NE) with the hydrogel to obtain the TFB loaded NEG formulation and DCN loaded NEG formulation.
d) Evaluating the prepared NEG formulations for pH, viscosity, spreadability, drug content, and in-vitro drug release studies.
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2. A method for optimizing the formulation of TFB (Trifluperidol) and DCN for topical application, comprising preparing a TFB and DCN loaded nanoemulsion gel (NEG) using Carbopol-934 and triethanolamine as claimed in claim 1.
3. The method for preparing Topical Nanoemulgel Formulation of Tofacitinib Citrate (TFB) and Diacerein (DCN) as claimed in claim 1, wherein Oleic acid, tween 80, and propylene glycol were selected as oil, surfactant, and co-surfactant, respectively.
4. The Topical Nanoemulgel Formulation of Tofacitinib Citrate and DiacereinMaximum prepared by a method claimed in claim 1, exhibits decrease in the value of paw volume of albino rats treated with DCN-NEG (1.257 ± 0.042 ml) in contrast to TFB-NEG (1.365 ± 0.082 ml) treated group.
5. The Topical Nanoemulgel Formulation of Tofacitinib Citrate and DiacereinMaximum prepared by a method claimed in claim 1, exhibits reduction in paw diameter of albino rats be more with TFB-NEG (0.682 ± 0.051 cm) treated group as compared to DCN-NEG (0.688 ± 0.029 cm).