DESC:FILED OF INVENTION: Synergistic Phytochemical blend in Anti-arthritic and chondrocytes regeneration potential and its process for preparation thereof. The present invention provides synergistic nutraceutical blend comprising Boswellia serrata extract selectively enriched in 3-O-acetyl-l l-keto-ß-boswellic acid (AKBA). It has good anti-inflammatory response in arthritis. Curcumin 95% demand also increased for its multiple benefits as an anti-infective, and an anti-inflammatory support. Satavari has good antioxidant and anti-inflammatory property, and its saponin content helps to improve solubility of Boswellic acids, AKBA and Curcumin itself. This innovative synergistic composition is proven to prevent, control and treat inflammation, pain and disease conditions related to or associated with Arthritis, Joint inflammation, Muscular inflammation and like that as per Anti-arthritic and chondrocytes regeneration study and clinical evidence. BACKGROUND OF INVENTION:
There is almost 100+ available source indicated arthritis and its related type chronic painful disease. Generally, 2 most common type called rheumatoid arthritis (RA) and osteoarthritis (OA). OA is more common in old age people than RA. Both involve inflammation in joints but RA Cause more inflammation. OA and RA have almost same symptoms and affected in multiple joints but one major different of RA is autoimmune and can be affected in multiple joints and OA is develop with more common in aged people and affect in few joints on one side. Osteoarthritis (OA) is the commonest form of inflammatory joint disease. Unfortunately, to date, there is no appropriate treatment for OA and RA. Boswellia serrata is considered as a potent anti-inflammatory, anti-arthritic and analgesic agent that may be a future of drug for arthritis. Based on our invention and clinical study, Boswellia and its extract in the form of 30% AKBA and Boswellic acid (60%) and Synergy form of 95% Curcumin and Saponin of Satavari has been effective and safe treatment option for patient with arthritis, and the recommended duration of treatment with Boswellia and its extract is at least 4 -12 weeks. As concentration of active ingredient (Phytochemical) increased, solubility and bioavailability decrease. Reported solubility enhancement of Boswellic acid, AKBA, Curcumin is available but it is by adding high number of excipients, with expensive and complex technology like SMEDDS contain poor yield of active. Here with we design Boswellic acid (60%), AKBA (30%), Curcumin (95%) and Satavari Saponin selected in ratio (70:20:5:5) for unique granulation. 2.5% of Saponin from Satavari used as a granulating agent in Aqueous medium for Granulation, and remaining 2.5% Saponin from Satavari is to be added during granulation simultaneously.
SUMMERY OF THE INVENTION: In arthritis, Osteoarthritis (OA) is an inflammatory joint disease that mainly damages articular cartilage. It is characterized by the degradation of articular cartilage and involving the entire joint tissue, which eventually leads to joint degeneration, fibrosis, fracture, defect and damage to the entire articular surface.
Epidemiological studies show that there are currently 355 million people with arthritis worldwide, and arthritis has become the world’s number one disabling disease. RA is autoimmune, Pain relief, Anti-inflammatory drug is not a solution for same but once it’s diagnosed at initial stage, then some evidence with Boswellia serrata and curcumin there about recovery and cell regenerate, but has limitation due to its poor characteristic i.e., solubility and bioavailability. Boswellia serrata, Curcumin is widely used in herbal, nutraceutical and supplements industry but due to its drawback it’s difficult to convert in precious dose.
Composition contains synergy blend of Boswellic acid (60%), AKBA (30%), Curcumin (95%) and Satavari Saponin prepared with novel wet granulation technique with the help of Satavari saponin as a granulating agent. It helps to improve solubility and bioavailability of BA, AKBA and Curcumin simultaneously. Study shows linearly improvement of solubility of Synergy blend. Curcumin also demonstrated its good solubility with synergy ingredients. There is also found tremendous improvement in micromeritics property of synergy blend, hence helpful to minimize other additives or excipients in finished dosage. Synergy blend converted in to tablet dosage form and release profile set up to 100% release in 45 minutes. It is made without diluents, binder or other dosage modifying agent. Optimize dose evaluated for further physical evaluation and stability study. DETAILED DESCRIPTION OF INVENTION:
Osteoarthritis (OA) is a debilitating disease, which primarily affects the hips and knees, the body-weight-bearing joints. Breakdown of the extracellular matrix of articular cartilage by the proinflammatory cytokine-induced tissue proteinases is the hallmark feature of the pathophysiology of OA. The clinical presentations of OA are pain and degenerative changes in the tissues surrounding the affected joints. Globally, OA of hip and knee is the 11th highest contributor to the disability with enormous economic burden. Some of the important factors that induce the progression of OA are chronic inflammation and gradual structural changes/structural remodeling within the joint tissues.
Boswellic acid is the active ingredient in Boswellia serrata; it has shown significant pharmacological activity in the treatment of inflammatory diseases such as rheumatoid arthritis, chronic bronchitis, asthma and chronic inflammatory bowel diseases (ulcerative colitis and Crohn’s disease). Current research showed that 3-O-Acetyl-11-keto-beta-boswellic acid (AKBA) is the one boswellic acid with strong pharmacological activity. Curcumin is a poorly water-soluble or insoluble phytochemical and its oral bioavailability is very low. Novel granulation technology has been successfully developed to improve the solubility and oral absorption of curcumin. It improves solubility, Photostability and Micromeritics property also. Final optimized synergy complex is selected for statistical analysis to optimize tablet dosage form gives complete release in 45 min at highest bioavailability.
Example: Table 1: Synergy blend of Phytochemicals for Anti Inflammatory
Botanical name of Plant
Active constituents
Of Synergy blend
Active Concentration
Ratio (%)
Boswellia serrata
Boswellic acid
60%
70
Boswellia serrata
AKBA
30%
20
Curcuma Longa
Curcumin
95%
5
Asparagus
Saponin
10%
5
Below are the details for solubility enhancement with the help of granulating agent are as follow: Table 2: Solubility Enhancement
Active constituents
Of Synergy blend
Granulating agent
% Improvement in solubility
% Improvement in Bioavailability
Boswellic acid 60%
Saponin 2-5%
39%
Study Pending
AKBA 30%
Saponin 2-5%
24 %
Study Pending
Curcumin 95%
Saponin 4-6%
66%
Study Pending
AKBA (30%), Boswellic acid (60%) and Curcumin (95%) are active constituents of synergy blend, useful for prevention, control and treatment of arthritis disease but shows poor characterises (solubility and bioavailability). Saponin content as granulating agent helps to improves solubility of BA, AKBA and Curcumin. Table 3: % Cumulative release comparison of synergy blend with other products
Time (min)
Synergic Blend
Marketed product 1
Marketed Product 2
0
0.00%
0.00%
0.00%
15
37.98%
11.09%
19.09%
30
78.98%
24.21%
31.05%
45
99.97%
33.45%
45.32%
Without use of diluents, binder or any other dosage modifying agent, release profile is set to 100% in 45min.
Details of procedure for product preparation and range of excipients.
Table 4: Following are the steps of procedure for product preparation
Excipient (Natural Saponin)
Loading time
Heating time (50-65 0C)
Asparagus Saponin (2-5%w/v)
9-24 hr
30-60 min
Table 5: Granulation process parameter to produce synergic blend
Kneading time (min)
Drying (45-600C)
Pulverize Through
30-60 min
45-90 min
22 and 60
Steps:-
1. Individual Phytoconstituent are cross check by suitable analytical technique (DSC, IR, UV).
2. Synergic blends are proceed for mixing via solid dispersion method. Composition contain synergic blend of Boswellic acid (60%), AKBA (30%) and Curcumin (95%) are mixed for 30-60min.
3. Expect Satavari (Saponin) individual extract reprocesses with granulation as a granulating agent to improve micromeritics property and enhance phytochemical characteristic.
4. Blend of phytoconstituents are subjected for drying at 45-600C for 45-90min using tray dryer.
5. Then, proceed for pulverization at initially sieve number 22 and then 60.
6. Lastly, check flowability of synergic blend as per free flow compliance.
As the formulation is excipientless, individual active constituent replaces the role of excipient. Table 6.1 of 6: Excipient and range
Botanical name of Plant
Active constituents
Of Synergy blend
Active Concentration
Ratio (%)
Boswellia serrata
Boswellic acid
60%
70
Boswellia serrata
AKBA
30%
20
Curcuma Longa
Curcumin
95%
5
Asparagus
Saponin
10%
5
Table 6.2 of 6: Details of Excipients are role of it to make product better.
Excipient
The Patent Role
Boswellic acid
Good inflammatory Response in arthritis
AKBA
Good inflammatory Response in arthritis
Curcumin
Anti-infective and anti-inflammatory support
Saponin Good antioxidant and anti-inflammatory property, Saponin content helps to improve solubility
o Boswellic acid is the active ingredient in Boswellia serrata; it has shown significant pharmacological activity in the treatment of inflammatory diseases such as rheumatoid arthritis, chronic bronchitis, asthma and chronic inflammatory bowel diseases (ulcerative colitis and Crohn’s disease).
o Current research showed that 3-O-Acetyl-11- keto-beta-boswellic acid (AKBA) is the one boswellic acid with strong pharmacological activity.
o Curcumin is a poorly water-soluble or insoluble phytochemical and its oral bioavailability is very low. Novel granulation technology has been successfully developed to improve the solubility and oral absorption of Curcumin. It improves solubility, Photostability and Micromeritics property also.
o Final optimized synergy complex is selected for statistical analysis to optimize tablet dosage form gives complete release in 45 min at highest bioavailability.
Table 7: Excipients category and range
As the formulation is excipientless, individual active constituent replaces the role of excipient. Excipients Concentration (%)
Ratio (%)
Boswellic acid
60%
70
AKBA
30%
20
Curcumin
95%
5
Saponin
10%
5 Table 8: Stability study till 3 months
Sr. No.
Parameters
Conditions
Initial
40°C & 75% RH
40°C & 75% RH
40°C & 75% RH
0 Day
1 Month
2 Months
3 Months
1
Disintegration Time
90-120sec
90-120sec
90-120sec
90-120sec
2
%Release in 45 min
99.98
99.90
98.00
98.13
3
Weight variation
Less than 1%
Less than 1%
Less than 1%
Less than 1%
4
Friability
0.871
0.869
0.865
0.863
5
Hardness
4-5
4-5
4-5
4-5
No significant changes are notified while stability study upto 3months. Data available are in compliance with pharmacopeia.
Table 9: Preformulation study of synergic blend
Micromeritics properties
Phytochemical
Pharmacopeial standard
Synergy blend
Pharmacopeial standard
Angle of Repose
Boswellic acid 60%
Poor
With Saponin 2-5%
Good to Excellent
Carr’s index
AKBA 30%
Poor
With Saponin 2-5%
Good to Excellent
Hausaner Ratio
Curcumin 95%
Poor
With Saponin 4-6%
Good to Excellent
Rational why this product is showing better activity where all ingredients are known.
Purpose behind this invention is to convert excipientless precise dose made up without excipient, enhancing therapeutic action with elimination of side effects.
Screening of several phytoconstituents/phytochemicals leading to formation of statistically design synergic components with improved dosage evaluation parameters, therapeutically efficacy and potency.
Synergic blend is produced without adding chemical based preservative, excipients, additives, and dosage modifying agent executing synergic therapeutic effect.
Also, aqueous transparent coating developed by OEM technology results in improved post-formulation parameters.
GP life Joint Support tablet is safe and significantly effective in reducing pain, swelling, and stiffness of knee joints and improving mobility of knee joints in patients with OA as Compare to Placebo. There is 50 % significant reduction of elevated serum CRP levels in treatment group compared to 23.55 % in placebo group. The Uric acid in treatment group is reduced by 11.08% while in placebo group it is increased bt 1.23%. There is 48.84% significant reduction in treatment group compared to 14.75 % in placebo group. There is 74.70% significant reduction in treatment group compared to 5.65 % in placebo group. In the present study the mean age of male and female subjects in treatment and placebo group are comparable and ranged from 47.97 to 52.37 years. Table 10: CRP Analysis
CRP mg/L (Mean±SD)
Duration
Test
Placebo
P value
Baseline
4.42±2.66
4.50±2.85
0.865
Day 90
2.21±1.49
3.44±3.02
Mean diff (Baseline – Day 90)
2.21±1.37
1.06±0.69
<0.001
% Reduction
50%
23.55%
P value
<0.001
<0.001
Data analyzed by student t test. Significant at p<0.05
There is 50 % significant reduction of elevated serum CRP levels in treatment group compared to 23.55 % in placebo group.
Table 11: Uric acid Analysis
Uric Acid mg/dl (Mean±SD)
Duration
Test
Placebo
P value
Baseline
4.51±2.62
4.05±1.32
0.218
Day 90
4.01±1.44
4.10±1.12
Mean diff (Baseline – Day 90)
0.51±2.49
-0.05±0.53
0.0009
% Reduction
11.08%
-1.23%
P value
0.002
0.197
Analyzed by Wilcoxon signed rank test for within group analysis and Mann Whitney U test for between group analysis. Significant at p<0.05
The Uric acid in treatment group is reduced by 11.08% while in placebo group it is increased bt 1.23%. Table 12: IL-6P Analysis
IL-6 pg/mL (Mean±SD)
Duration
Test
Placebo
P value
Baseline
5.61±3.82
6.10±3.61
0.051
Day 90
2.87±1.35
5.20±3.78
Mean diff (Baseline – Day 90)
2.73±3.41
0.89±0.53
0.012
% Reduction
48.84%
14.75%
P value
<0.001
<0.001
Analyzed by Wilcoxon signed rank test for within group analysis and Mann Whitney U test for between group analysis. Significant at p<0.05
There is 48.84% significant reduction in treatment group compared to 14.75 % in placebo group.
Table 13: VAS Score Analysis
VAS Score (Mean±SD)
Duration
Test
Placebo
P value
Baseline
7.79±0.76
7.81±0.82
0.920
Day 90
3.80±0.77
7.37±0.49
Mean diff (Baseline – Day 90)
3.99±0.65
0.44±0.76
<0.001
% Reduction
51.21%
5.63%
P value
<0.001
<0.001
Analyzed by Wilcoxon signed rank test for within group analysis and Mann Whitney U test for between group analysis. Significant at p<0.05. There is 51.21% significant reduction in treatment group compared to 5.63 % in placebo group.
Table 14: WOMAC Score Analysis
WOMAC Score (Mean±SD)
Duration
Test
Placebo
P value
Baseline
66.73±4.64
62.24±3.42
<0.001
Day 90
16.88±2.76
58.72±4.14
Mean diff (Baseline – Day 90)
49.85±4.74
3.52±5.24
<0.001
% Reduction
74.70%
5.65%
P value
<0.001
<0.001
Data analyzed by student t test. Significant at p<0.05
There is 74.70% significant reduction in treatment group compared to 5.65 % in placebo group.
Table 15: Demographic details
Parameter
Treatment
Placebo
Group
Male (n=20)
Female(n=55)
Male(n=24)
Female(n=51)
Age (years)
53.85±9.37
51.84±9.69
46.88±4.86
48.49±6.73
Total Age (years)
52.37±9.58
47.97±6.20
In the present study the mean age of male and female subjects in treatment and placebo group are comparable and ranged from 47.97 to 52.37 years.
• SUMMARY
? Joint Swelling is found to be reduced from day 7th only.
? Reduction in inflammatory parameters suggests the anti-inflammatory and anti-arthritic action of Gplife Joint Support Product
? TNF-a levels decreased by 55% in the Gplife Joint Support Product group when compared with the Disease Control group.
? LT-BT4 levels decreased by 57% in the Gplife Joint Support Product group when compared with the Disease Control group.
? CRP levels decreased by 69% in the Gplife Joint Support Product group when compared with the Disease Control group.
? MMP-13levels decreased by 88% in the Gplife Joint Support Product group when compared with the Disease Control group.
? COMP levels decreased by 37% in the Gplife Joint Support Product group when compared with the Disease Control group.
? IL-6 levels decreased by 61% in the Gplife Joint Support Product group when compared with the Disease Control group.
? COMP might therefore be involved in the regeneration efforts of cartilage tissue as a factor secreted by chondrocytes to ameliorate the matrix breakdown. Gplife Joint Support Product can improve the chondrocyte production for the repair of damaged cartilage
? In this study, we have found that a decrease in MMP-13 stabilizes their extracellular matrix, impaired chondrocyte terminal maturation and enhanced their viability, and inhibited multiple effectors and regulators of chondrocyte differentiation. The decreased levels of MMP-13 indicate the chondroprotective effect and anti-arthritic potential of the Gplife
? joint support product. Gplife Joint Support Product is a potent inhibitor of the production of inflammatory and catabolic mediators by chondrocytes, suggesting that this natural treatment could be efficient in the treatment of osteoarthritis
? The histopathology of joints in the present study suggests that the damage to the cartilage and subchondral region in the Gplife Joint Support Product is not as severe as in disease control because the damage happed by an inducer is found to be recovered in Gplife Joint Support Product group. This indicates that Gplife Joint Support Product has chondroprotective and crondroregeneration activity
• Evaluation of Joint Thickness/Swelling
The knee swelling is calculated using the Vernier caliper scale.
In the disease control group, on Day 28, the animals in the diseased group showed a reduction in activity as represented by fewer distance traveled and higher immobility time. Meanwhile, it is found that the amount of distance traveled in those treated with Gplife Joint Support product is significantly higher than those treated with disease control. The fact that the immobility time of animals treated with Gplife Joint Support product is lower than that of control patients is also found to be significant.
TNF-a levels decreased by 55% in the Gplife Joint Support Product group when compared with the
Disease Control group.
There is a significant increase in TNF-a levels in the Disease Control group compared to the Normal Control group. The level of TNF-a is significantly low in the STD group and Gplife Joint Support Product group when compared to the Disease Control group.
LT-BT4 levels decreased by 57% in the Gplife Joint Support Product group when compared with the Disease Control group.
There is a significant increase in LT-BT4 levels in the Disease Control group compared to the Normal Control group. The level of LT-BT4 is significantly low in the STD group and Gplife Joint Support Product group when compared to the Disease Control group.
Evaluation of Level of CRP (ng/ml) after 28 days
CRP levels decreased by 69% in the Gplife Joint Support Product group when compared with the Disease Control group.
There is a significant increase in CRP levels in the Disease Control group compared to the Normal Control group. The level of CRP is significantly low in the STD group and Gplife Joint Support Product group when compared to the Disease Control group.
MMP-13levels decreased by 88% in the Gplife Joint Support Product group when compared with the Disease Control group.
There is a significant increase in MMP-13levels in the Disease Control group compared to the Normal Control group. The level of MMP-13is significantly low in the STD group and Gplife Joint Support Product group when compared to the Disease Control group.
COMP levels decreased by 37% in the Gplife Joint Support Product group when compared with the Disease Control group.
There is a significant increase in COMP levels in the Disease Control group compared to the Normal Control group. The level of COMP is significantly low in the STD group and Gplife Joint Support Product group when compared to the Disease Control group.
The present study found that the biomarkers COMP and MMP-13 are significantly lower in the Gplife joint support product. MMP-13 is a key enzyme in osteoarthritis that plays a central role in the degradation of articular cartilage. Its degree of expression is increased in animals with osteoarthritis and thus, inhibition of MMP-13 might be a new therapeutic target.
MMP 13 levels are significantly decreased in the Gplife joint support product group when compared to the disease control. This suggests that the Gplife joint support product showed anti-arthritic properties, as well as crondroregeneration properties.
HISTOPATHOLOGY EVALUATION
Microscopic examination of the knee joint and bone tissue of rats from the normal control group did not reveal any lesion of pathological significance. Rats of the disease control group showed enlarged synovial lining cell layer, synovial hyperplasia, increased synovial vascularity, infiltration of inflammatory cells, pannus formation, cartilage erosion, and bone erosion. To conclude or for interpreting the effect of treatment, it has been suggested to correlate other study blood parameter data.
RADIOLOGICAL EVALUATIONS OF HIP AND KNEE JOINTS OF ARTHRITIS IN RATS
METHODOLOGY
Digital X-Ray images of hip and knee joints are processed for radiological evaluations. The following parameters are considered for radiological evaluations of the bone and joint:
1) Swelling of the soft tissue around the joints
2) Periosteal reaction/hypertrophy
3) Narrowing of the joint space
4) Periarticular osteoporosis
5) Bone destruction/erosions
6) Any other lesions
• Grading of abnormalities is done according to the following severity level:
• Normal- 0, Slight- 1, Moderate-2, Severe-3
Table 16: Individual Animal Radiology Observations
Group
Joint
Swelling of the soft tissue around the joints
Periosteal reaction
Joint space narrowing
Bone erosion
Osteoporosis
Normal Control
Knee
0
0
0
0
0
Disease Control
Knee
3
3
2
2
2
Standard
Knee
1
1
2
1
1
Treatment- 100
Knee
1
0
2
1
2
Treatment- 200
Knee
1
1
1
2
1
? Note: Individual animal radiology score is given.
? Severity score: 0= Normal, 1= Slight, 2= Moderate, 3= Severe ,CLAIMS:A Synergistic nutraceutical blend in Anti-arthritic and chondrocytes regeneration potential and its process for preparation thereof. 2. The synergistic nutraceutical blend as claimed in claim 1, wherein the synergistic nutraceutical blend comprising Boswellia serrata extract selectively enriched in 3-O-acetyl-l l-keto-ß-boswellic acid (AKBA), curcumin and shatavari. 3. The synergistic nutraceutical blend as claimed in claim 1, wherein the synergistic composition is proven to prevent, control and treat inflammation and pain, disease conditions related to or associated with Arthritis, Joint inflammation, Muscular inflammation