Synergistic Composition

Abstract

The present invention relates to novel compositions comprising at least two different omega-3 fatty acid sources, including 5% (w / w) surf clam eggs in powder or oil form and green lipped mussel in powder or oil form. The compositions are particularly useful as / in supplements and / or animal or human foods. The invention may further relate to the maintenance or treatment of human and veterinary conditions, such as use in ameliorating inflammation or joint-related disorders that may be associated with inflammation.

Description

[Technical Field]

[0001] The present invention relates to novel compositions comprising a combination of two different blended marine-derived ingredients, with at least two ingredients comprising at least hoki roe (HR) powder or oil, preferably in combination with green-lipped mussel (GLM) powder or oil.

[0002] The invention further extends to the use of this composition in human and veterinary applications, particularly in supplements or nutraceuticals, thereby providing nutritional support for health, particularly joint health. [Background technology]

[0003] Nutraceuticals, foods, and dietary supplements are often used to improve or support joint structure and maintain joint mobility in mammals, including humans and animals.

[0004] Biological inflammatory pathways are directly linked to joint health, including, for example, joint mobility. Potentially, lipids, including inflammation-resolving lipid mediators, and bioactive peptides may also have a role, but may act through different pathways.

[0005] The effects have been shown to be derived from essential fatty acids (EFAs), and several enzymes associated with biological inflammatory pathways appear to be affected by the presence or absence of these EFAs, and also depend on the type of EFA.

[0006] High levels of omega (ω) polyunsaturated fatty acids, especially ω-3, are likely to support the body's natural anti-inflammatory processes and may therefore be beneficial. In particular, these EFAs are metabolized into anti-inflammatory eicosanoids. The presence of ω-3s reinforces existing anti-inflammatory pathways because they are preferentially acted upon in this process, whereas in their absence, natural substrates, including other EFAs such as arachidonic acid, are metabolized to produce pro-inflammatory substances.

[0007] Although GLM powder can be used to provide a source of omega-3 in supplement compositions, the quality and content of omega-3 fatty acids is not always reliable and may not always be sufficient for products intended to maintain joint health in humans or animals.

[0008] However, it remains desirable to provide compositions that are effective for use in supporting joint health, whether as long-term maintenance products, e.g., foods or dietary supplements, or as short-term dietary solutions. In particular, it is becoming increasingly interesting to be able to provide a high-specification GLM powder source that correlates with the desired technical specifications sufficient to meet these applications. Summary of the Invention

[0009] The present invention therefore arose out of the continuing need to provide new compositions that are useful in the commercial applications described above. The present invention relates to a composition comprising a combination of two powder or oil components, including a green lipped mussel (GLM) powder or oil component and at least 5% (w / w) of surf clam roe (HR) powder or oil component.

[0010] The present invention also relates to a composition comprising a blend of at least two different powder or oil components, each comprising omega-3 fatty acids, wherein a first of the at least two different powder or oil components comprises 5% (w / w) or more HR powder or oil, and the blend of the at least two different powder or oil components together provides at least 3.3% (w / w) total omega-3 fatty acids in the composition. Preferably, the second of the at least two powder components is a GLM powder or oil.

[0011] First, the applicant has observed that the selection of the source of omega-3 fatty acids is very important when providing a composition for the aforementioned uses. The omega-3 fatty acids in any commercially available powder source vary widely. Preliminary studies by the applicant have confirmed that the specifications of some GLM powders do not provide enough omega-3 to meet the technical standards for the intended purpose, compared to higher specification products that are rare and less readily available on the market.

[0012] However, GLM, even at low levels, remains an important basic omega-3 fatty acid source, and the inventors further investigated other ways in which a GLM-based composition would still provide a solution. They found that the inclusion of a minimal amount of another specifically selected omega-3 fatty acid source in combination with a lower specification GLM powder raised the omega-3 content to an acceptable level. Through testing, they found that this solution could be achieved by blending at least 5% HR powder with a lower specification GLM powder.

[0013] It has further been found through in vitro assay testing that by incorporating specific selections or combinations of omega-3 ingredients, unexpectedly enhanced levels of inhibitory or modulating activity can be observed in the composition. Such activity is strongly related to supporting inflammatory response pathways and maintaining optimal joint health. The anti-inflammatory activity of the blend of HR powder and GLM powder is preferably measured as a coefficient of 0.05 to 0.25, as determined according to the method described herein below.

[0014] The Applicant has further been able to demonstrate that, if the source remains the same, it is possible to use the HR and / or GLM components in the form of oils to obtain a composition with the same technical effect, thereby providing a further solution to the problem based on the basic concept of the present invention.

[0015] In similar assays as performed with the powder component, the effect of this particular two-oil component combination on inflammatory pathways is greater than would be expected from the sum of the individual component parts. Thus, the combination of these two ingredients, whether in powder or oil form, is better at inhibiting or regulating inflammation levels as measured by relative COX2 activity than would be predicted from the levels disclosed in the art. In other words, Applicant has demonstrated that whether the combination of ingredients is selected from powder or oil form, there is a synergistic effect that is useful for maintaining a better level of support for the anti-inflammatory system.

[0016] The in vitro inhibitory or modulating activity observed by the applicant appears to be far greater than the simple additive effect expected from the combination of omega-3 EFA levels alone. Accordingly, the applicant has determined that a most unexpected synergistic effect is achievable by the present invention. That is, by specifically incorporating HR at de minimus levels, and / or further specifically selecting this powder or oil component in conjunction with a lower specification GLM extract, a higher level of competitive enzyme activity is maintained than would be expected based on omega-3 EFA content alone. In an embodiment, the present invention therefore relates to a food product, supplement, or nutraceutical comprising a composition having the ingredients recited in the claims and described herein.

[0017] Such compositions meet or exceed required technical standards in terms of content and / or activity, thus enabling consistent and improved efficacy of GLM-based products for the purpose of supporting joint health. The present invention is useful for providing foods or supplements containing the compositions described herein. The foods or supplements may further contain one or more useful additives that are desirable to make the supplement more palatable or may provide other benefits. The additives may include one or more additives including, but not limited to, glucosamine, hyaluronic acid, and chondroitin sulfate.

[0018] In preferred embodiments of either aspect of the invention, the HR powder or oil is in the range of 5% to 95% (w / w) of the total composition mass. In embodiments, the HR powder or oil component may comprise a total fat content of 14% to 40% (w / w), but importantly, a total omega-3 fatty acid content of 2% to 20% (w / w).

[0019] With respect to GLM content, the GLM powder or oil comprises a total fat content of 7% to 13% (w / w). In embodiments, it is contemplated that the GLM powder or oil comprises a total omega-3 fatty acid content in the range of 2.0% to 7.0% (w / w), ideally 2.0% to 5.0% (w / w).

[0020] The compositions of the present invention are particularly useful as human supplements or veterinary products, and can be used to prevent, maintain low levels of, and / or treat inflammation. The compositions themselves and / or foods or supplements containing the compositions can be used to treat and / or maintain health, preferably joint health, in humans or animals. Furthermore, treatment and / or maintenance includes supporting anti-inflammatory pathways in humans or animals.

[0021] The present invention further extends to a method for maintaining joint health comprising administering a composition disclosed herein to an animal in need thereof, in an embodiment, maintaining health comprises supporting anti-inflammatory pathways in the animal, including preventing inflammation.

[0022] The present invention extends to a method of treating a subject, the method comprising administering to the subject a composition disclosed herein, the subject being a human or an animal. In embodiments, the method treats inflammation or maintains low levels of inflammation in the subject, the subject being an animal. In embodiments, the method maintains the health or joint health of the subject. [Brief explanation of the drawings]

[0023] [Figure 1] FIG. 1 is a graph showing the dose-response curve for the gold standard GLM. [Figure 2] FIG. 2 is a graph showing the testing of samples with varying GLM quality. [Figure 3] FIG. 3 is a graph showing the omega-3 / COX2 ratio data. [Figure 4] FIG. 4 is a graph showing a screening test of marine-derived samples comparing GLM, abalone, Mytilus edulis, and HR. [Figure 5] FIG. 5 is a graph showing the effect of specific % combinations showing predicted and actual effects on the inflammatory process. [Figure 6] FIG. 6 is a graph showing the concentration-dependent inhibition or modulation of COX2 of three different sample powders. [Figure 7] FIG. 7 is a graph showing the anti-inflammatory activity versus powder blend % compared to the expected trend lines for H powder and X powder. [Figure 8] FIG. 8 is a graph showing the anti-inflammatory activity of the x+H blend relative to total omega-3 fatty acid levels compared to the expected trend lines for H and X powders. [Figure 9]FIG. 9 is a graph showing that combinations of these two ingredients in oil form (at various blend percentages) are more effective at modulating COX2 activity than predicted by the trend line. DETAILED DESCRIPTION OF THE INVENTION

[0024] GLM is used to provide EFAs to the body. They support the anti-inflammatory process by being preferentially activated in the anti-inflammatory system, resulting in the production of anti-inflammatory mediators. In the absence of these omega-3 EFAs, other EFAs, especially arachidonic acid, are used as substrates, resulting in the production of pro-inflammatory mediators.

[0025] A dose response appears to be effective depending on the amount of GLM used, i.e., with increasing concentrations there is a decrease in the activity of the inflammatory process (COX2 activity), see Figure 1.

[0026] We found that when GLM was used at the same concentration, but crucially with a lower EFA specification, there was a reduced effect on the inflammatory process, see Figure 2.

[0027] Furthermore, the present inventors have found a correlation between omega-3 EFAs and the activity of the inflammatory process, i.e., with an increase in omega-3 EFAs, the activity of the inflammatory process decreases, see Figure 3.

[0028] Additionally, certain marine species were found to be more effective at reducing the level of inflammatory processes despite comparable total EFA levels, suggesting that bioactivity is associated with specific EFAs compared to all tested (see Figure 4).

[0029] HR has been identified as a fatty acid source that appears to have a higher profile for certain EFAs and is equivalent to GLM for some EFAs. Therefore, the possibility of using HR as a replacement / supplement for low-EFA GLM and low-concentration full-spec GLM was further investigated.

[0030] Based on the relative effects of GLM and HR on the inflammatory process, several combinations of low-concentration GLM / HR or low-EFA GLM / HR were tested. With the goal of maintaining the expected effect from full-concentration, full-specification GLM, the expected additive effect was calculated based on the inflammatory performance of the original GLM and HR samples. See Figure 5.

[0031] Indeed, the inventors have found that the effect of the combination on the inflammatory process is greater than would be expected from the sum of the individual component parts. The present invention demonstrates a clear synergistic effect that can be used to maintain a better level of support for the anti-inflammatory system.

[0032] As previously mentioned herein, EFAs are considered important components of any composition intended to help support the natural anti-inflammatory processes associated with the joints of humans or animals.

[0033] In particular, omega-3 EFAs appear to act as dual competitors for arachidonic acid oxidation by both the cyclooxygenase (COX) and lipoxygenase (LOX) pathways and are therefore useful in the above applications.

[0034] EFAs and omega-3 fatty acids can be found, inter alia, in green lipped mussels (GLM) or green shell mussels (GSM; Perna canaliculus). As previously mentioned, because specifications for GLM powder vary, and because many existing powder sources of GLM alone do not necessarily provide sufficient omega-3 fatty acid content, research was conducted to find alternatives or supplements.

[0035] Various marine-derived sources were reviewed as potential sources of desirable omega-3 fatty acid content. Generally, marine-derived sources of high omega-3 fatty acids are oils from shellfish or fish organs. These various types of oils were screened as potential replacements for some of the components that make up GLM in order to improve the overall omega-3 fatty acid content. Although good sources of omega-3 fatty acids in themselves, it proved difficult to blend the oils with GLM powder to obtain a uniform powder suitable for formulation into applicable products.

[0036] However, one such source of ω-3 fatty acids was dried egg powder from the whitefish Macruronus novaezelandiae (Hoki), or hoki (hoki).Hoki egg (HR) powder was tested to determine whether it would improve the EFA content, especially the ω-3 fatty acid content, when added to GLM powder.

[0037] The suitability of the HR powder was determined by measuring the levels of omega-3 fatty acids in the GLM powder before and then after the addition of the HR powder to determine whether the combination raised the omega-3 levels to the required specifications, which were achieved with GLM powder, which has a relatively high omega-3 fatty acid specification known to produce commercially desirable therapeutic results.

[0038] To confirm that the novel blend of powders also exhibited the desired anti-inflammatory activity (despite the inclusion of low omega-3 content GLM), in vitro enzymatic activity was tested and compared to the standard desired specification product used by the applicants.

[0039] A commonly used in vitro process for measuring anti-inflammatory activity was used, namely, by determining the level of in vitro competition of the COX2 enzyme, a pathway known to be strongly involved in inflammatory processes as previously discussed herein.

[0040] method New Zealand green lipped mussel (GLM) powder and New Zealand HR powder were obtained from commercial sources, and lipid extracts were obtained from individual powders or blends of powders using the Bligh and Dyer method, as described in the following published reference (Bligh and Dyer, A rapid method of total lipid extraction and purification. Canadian Journal of Biochemistry and Physiology 1959, 37: 911-917). The fatty acid composition of the lipid extracts was determined using AOAC method 963.22.

[0041] result The first data set was generated by measuring and comparing the amounts of omega-3 fatty acids in a standard reference source GLM product (X) known by the applicant to have acceptable specifications and therefore the appropriate and desired omega-3 fatty acid requirements for use. A further source GLM powder (x) known to have a lower specification was also measured.

[0042] GLM x powder alone gave poor results, with significantly lower omega-3 fatty acids measured than the reference powder GLM(X), along with lower COX2 activity or regulation, and therefore below the required specifications for commercially desirable applications as previously described herein.

[0043] Novel combinations were tested to determine how omega-3 fatty acid levels are affected when low specification product x is supplemented with amounts of different powders known to be rich in omega-3s.

[0044] HR powder (H) was previously selected from various sources based on its good potential to enhance total omega-3 fatty acid content and its ease of blending with GLM powder. To determine whether a low specification GLM powder could be raised to the required specification by the addition of an HR powder, it was determined that, theoretically, the addition of at least 5% (wt / wt), and preferably 8% (wt / wt), of a specific HR powder (BN:HR114) to a low specification GLM powder (x) would raise the total omega-3 content score (a measure of the % fatty acids in the total fat of the product) to a level that met the required specification.

[0045] As shown in Table 1, measurements determined that the inclusion of 8% (wt / wt) HR powder (H) in a blend with low specification GLM powder (x) increased the omega-3 fatty acid content to 3.5% (wt / wt), very close to the level exhibited by standard powder X (BN32132).

[0046] [Table 1]

[0047] Biological Activity and Graphical Analysis Different quality specifications of GLM (with different levels of omega-3 fatty acids) were combined with HR powder in different ratios. The total omega-3 fatty acids of the combinations were then measured, and the COX inhibition or modulation scores of these powders were compared with the source GLM powder (x) and hoki powder (H).

[0048] The anti-inflammatory activity of the above examples was then measured to identify promising potential biological effects of the combinations compared to predicted activity based on novel combinations of omega-3 fatty acids from source sources.

[0049] COX2 inhibition or modulation was determined by incubating lipid extracts of the powder blend combination with commercially available mammalian COX2 enzyme, and enzyme activity was measured as the co-oxidation of N,N,N,N'-tetramethyl-p-phenylenediamine (TMPD) by prostaglandin G2 (PGG2), which is blue in color and easily measurable at 611 nm.

[0050] The COX2 inhibitory or modulating activity of the powder blend combination was determined from lipid extract concentration-dependent inhibition or modulation of the COX2 enzyme or from the concentration of lipid extract required to inhibit or modulate the COX2 enzyme by 50% (IC 50 The powder samples were identified by their anti-inflammatory activity (AI) score, calculated by multiplying the inverse of the lipid mass index (μm) by the total weight of lipid extracted from the powder blend combination.

[0051] As can be seen, the concentration-dependent inhibition or modulation of COX2 by the low specification GLM powder (x) blended with 8% (wt / wt) HR powder (H) to form the x+H (BN32127) powder was determined to be significantly greater than that of GLM powder X (BN32132) alone, which exhibits comparable omega-3 fatty acid levels (as shown in Table 1) and is therefore assumed to be comparable.

[0052] Rather surprisingly, as specifically shown in Figure 6, the concentration-dependent inhibition or modulation of COX2 by x+H (BN32127) was also greater than that of 100% HR powder H (BN:HR114), which exhibited significantly higher omega-3 levels (as shown in Table 1) than x+H.

[0053] 7 further reveals that the anti-inflammatory (AI) activity exhibited by x+H at various H blend % (diamond data points) was greater than would be expected from the (AI) activity predicted by plotting a linear trend line representing the activity of GLM Powder X (left-most square data points) and HR Powder H alone (right-most data points). This linear trend line between the two represents what would be expected from a simple additive effect of the two powders, suggesting that the two, when combined, have an enhanced effect far beyond the sum of their individual effects.

[0054] This trend was further seen in Figure 8, as the anti-inflammatory (AI) activity (diamond data points) of x+H at various powder blend % was also greater than predicted from the total omega-3 fatty acid levels of H and X alone (as shown in Figure 3).

[0055] The large square data points exhibit a linear trendline relationship, indicating the effect expected from a simple additive effect achieved by combining the two powders. However, as can be seen, the blended powder vastly exceeds the expected AI activity, indicating that this novel combination provides an enhanced effect, and therefore a synergistic effect, rather than a simple cumulative or additive effect.

[0056] We also tested oil-based combinations of these ingredients and found that the anti-inflammatory activity (circular data points) exhibited by various H blend percentages was also greater than would be expected from the sum of the oil component activities, confirming that oil-based component combinations are also effective in generating enhanced biological activity compared to the individual components.

[0057] Table 2 below shows the results of the oil-based ingredient testing.

[0058] [Table 2]

[0059] Figure 9 illustrates the implications of these results (shown in Table 2) in graphical form by plotting the relative COX2 activity that might be expected from 100% GLM oil (left-most square) and 100% HR oil (right-most square), each used alone. The linear trend line between the two plotted square data points shows the effect that might be expected from the simple additive effect of the two oils.

[0060] A plot of the results from the actual activity observed for various combinations (plotted as circular data points), also found in Table 2 above, suggests that the combination of oil components also has an enhanced synergistic effect beyond the additive effect of the individual oil components.

Claims

1. Green-lipped mussel (GLM) powder or oil, and at least 5% (w / w) of surfacing egg (HR) powder or oil, a combination of at least two different powder components or two oil components, A composition that synergistically inhibits COX2 activity.

2. 10. The composition of claim 1, wherein said composition of said at least two different powder or oil components together provides at least 3.3% total omega-3 fatty acids in said composition.

3. 3. The composition of claim 1 or 2, wherein the HR powder or oil is present in the range of 5% to 95% (w / w) of the total composition.

4. 4. The composition of any one of claims 1 to 3, wherein the GLM powder or oil has a total fat content of 7% to 13% (w / w).

5. 5. The composition of any one of claims 1 to 4, wherein the GLM powder or oil comprises a total omega-3 fatty acid content in the range of 2.0% to 5.0% (w / w).

6. 6. The composition of any one of claims 1 to 5, wherein the HR powder or oil comprises a total fat content of 14% to 40% (w / w).

7. 7. The composition of any one of claims 1 to 6, wherein the HR powder or oil comprises a total omega-3 fatty acid content of 2% to 20% (w / w).

8. A food, nutraceutical, or supplement comprising the composition of any one of claims 1 to 7.

9. 9. The food, nutraceutical, or supplement of claim 8, further comprising one or more additives selected from glucosamine and / or hyaluronic acid.

10. 10. A composition, food, nutraceutical or supplement according to any one of claims 1 to 9 for use in preventing or treating inflammation or maintaining inflammation at a low level in a human or animal.

11. 11. A composition, food product, nutraceutical or supplement according to any one of claims 1 to 10 for use in maintaining joint health in humans or animals.

12. 12. The composition, food, nutraceutical, or supplement of claim 10 or 11, wherein the preventative, therapeutic, or maintenance use comprises supporting anti-inflammatory pathways in humans or animals.

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