# US 2014/0193369 A1 · Anti-inflammatory food product

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- Number: US 2014/0193369 A1
- Document ID: US20140193369A1
- Status: Published application
- Kind: A1
- Application number: 13/735,547
- Filed: 2013-01-07
- Published: 2014-07-10
- Earliest claimed: 2013-01-07
- Assignees: WhiteWave Services, Inc.
- Applicants: Whitewave Services, Inc.
- Inventors: Neal Allan Bringe
- Web page: https://patentgrep.com/patent/US20140193369A1

## CPC

- C12Q 1/025: {for testing or evaluating the effect of chemical or biological compounds, e.g. drugs, cosmetics (antimicrobial activity C12Q1/18)}
- G01N 33/573: for enzymes or isoenzymes
- G01N 33/6863: {Cytokines, i.e. immune system proteins modifying a biological response such as cell growth proliferation or differentiation, e.g. TNF, CNF, GM-CSF, lymphotoxin, MIF or their receptors}
- G01N 33/6869: {Interleukin}
- G01N 33/88: involving prostaglandins {or their receptors}
- G01N 2333/525: Tumor necrosis factor [TNF]
- G01N 2333/5412: IL-6
- G01N 2333/545: IL-1
- G01N 2333/90254: Nitric-oxide synthase (NOS; 1.14.13.39)
- G01N 2800/7095: Inflammation
- C12Q: MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS (immunoassay G01N33/53); COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- G01N: INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES (measuring or testing apparatus or processes other than immunoassay, involving enzymes or microorganisms C12M, C12Q)

## Abstract

According to some embodiments, a method selects one or more inflammatory markers to evaluate. The one or more inflammatory markers selected from the group consisting of iNOS, COX-2, TNF-alpha, NO, PGE2, IL-6, and IL-1β. The method determines a first expression level and a second expression level of the one or more inflammatory markers. The first expression level is associated with a positive control sample comprising a type of human macrophages. The second expression level is associated with a test sample obtained by exposing a food product to a digest, the type of human macrophages, and a pro-inflammatory compound. The method performs a comparison between the first expression level and the second expression level and designates the food product according to an anti-inflammatory standard based on the comparison.

## Claims

### Claim 1

A method comprising:
obtaining a whole food product comprising a mixture of at least two ingredients selected from the group consisting of: a vegetable ingredient, a fruit ingredient, a seed, an herb, a spice, a plant-based protein ingredient, and a yogurt culture, wherein the mixture has been pasteurized and homogenized during a commercial process to form the whole food product;
obtaining a test sample according to steps comprising:
heating the whole food product to approximately 37° C.;
exposing the heated whole food product to digestive enzymes for between approximately 1 to 3 hours to form a digest;
obtaining an extract comprising a desalted supernatant of the digest; and
exposing the extract to RAW 264.7 macrophages and a pro-inflammatory compound for a pre-determined amount of time between approximately 20 and 30 hours;
obtaining a positive control sample that includes the RAW 264.7 macrophages and excludes the pro-inflammatory compound and the whole food product;
determining a first expression level of a PGE2 inflammatory marker, the first expression level associated with the positive control sample;
determining a second expression level of the PGE2 inflammatory marker, the second expression level associated with the test sample;
performing a comparison between the first expression level and the second expression level; and
designating the whole food product according to an anti-inflammatory standard, wherein the anti-inflammatory standard indicates to designate the whole food product as anti-inflammatory if the comparison between the first expression level and the second expression level yields a statistical p value less than 0.05.

### Claim 2 (depends on 1)

The method of claim 1, further comprising:
determining a third expression level of the one or more inflammatory markers, the third expression level associated with a negative control sample comprising the RAW 264.7 macrophages and the pro-inflammatory compound, the negative control sample excluding the whole food product;
wherein performing the comparison further comprises comparing the second expression level and the third expression level.

### Claim 3 (depends on 2)

The method of claim 2, wherein the anti-inflammatory standard indicates to designate the whole food product as anti-inflammatory if the second expression level exhibits at least a 66% reduction in PGE2 when compared to the third expression level.

### Claim 4 (depends on 2)

The method of claim 2, wherein the anti-inflammatory standard indicates to designate the whole food product as anti-inflammatory if:
the second expression level exhibits at least a 15% reduction in PGE2 when compared to the third expression level; and
expression levels of iNOS, COX-2, and TNF-alpha associated with the test sample exhibit at least a 15% reduction in each of iNOS, COX-2, and TNF-alpha when compared to respective expression levels of iNOS, COX-2, and TNF-alpha associated with the negative control sample.

### Claim 5 (depends on 2)

The method of claim 2, wherein the anti-inflammatory standard indicates to designate the whole food product as pro-inflammatory if the comparison between the second expression level and the third expression level yields a statistical p value less than 0.05.

### Claim 6 (depends on 1)

The method of claim 1, wherein the whole food product comprises a non-dairy milk.

### Claim 7 (depends on 1)

The method of claim 1, wherein the anti-inflammatory standard indicates to designate the whole food product as anti-inflammatory if expression levels of iNOS, COX-2, and TNF-alpha associated with the test sample each yield a statistical p value less than 0.05 when compared to respective expression levels of iNOS, COX-2, and TNF-alpha associated with the positive control sample.

### Claim 8 (depends on 1)

The method of claim 1, wherein the digestive enzymes comprises pepsin or pancreatin.

### Claim 9 (depends on 1)

The method of claim 1, wherein exposing the heated whole food product to the digestive enzymes comprises adding pepsin for 90 minutes at pH 2 followed by adding pancreatin for 90 minutes at pH 7.5.

### Claim 10 (depends on 9)

The method of claim 9, further comprising:
determining a third expression level of the one or more inflammatory markers, the third expression level associated with a negative control sample comprising the RAW 264.7 macrophages exposed to the pro-inflammatory compound for approximately the same pre-determined amount of time between approximately 20 and 30 hours, the negative control sample excluding the whole food product;
wherein performing the comparison further comprises comparing the second expression level and the third expression level.

### Claim 11

A method comprising:
selecting one or more inflammatory markers to evaluate, the one or more inflammatory markers selected from the group consisting of iNOS, COX-2, TNF-alpha, NO, PGE2, IL-6, and IL-1β;
determining an expression level of the one or more inflammatory markers associated with a test sample, the test sample comprising a food product exposed to a digest, a type of human macrophages, and a pro-inflammatory compound; and
designating the food product according to an anti-inflammatory standard based on the expression level of the one or more inflammatory markers.

### Claim 12 (depends on 11)

The method of claim 11, further comprising obtaining a negative control sample comprising the type of human macrophages and the pro-inflammatory compound; and
determining a second expression level of the one or more inflammatory markers, the second expression level associated with the negative control sample;
wherein the designating the food product according to an anti-inflammatory standard is based on comparing the expression level and the second expression level.

### Claim 13 (depends on 12)

The method of claim 12, the anti-inflammatory standard designates the food product as anti-inflammatory if the expression level for each of the selected inflammatory markers falls statistically below a corresponding predetermined threshold, the predetermined threshold indicating that the test sample expresses the selected inflammatory marker at a sufficiently lower level than the level at which the negative control sample expresses the selected inflammatory marker.

### Claim 14 (depends on 13)

The method of claim 13, wherein:
the predetermined threshold for iNOS is at least a 65% reduction compared to the second expression level;
the predetermined threshold for COX-2 is at least a 62% reduction compared to the second expression level;
the predetermined threshold for TNF-alpha is at least a 40% reduction compared to the second expression level;
the predetermined threshold for NO is at least a 19% reduction compared to the second expression level;
the predetermined threshold for PGE2 is at least a 66% reduction compared to the second expression level;
the predetermined threshold for IL-6 is at least a 24% reduction compared to the second expression level; and
the predetermined threshold for IL-1β is at least a 25% reduction compared to the second expression level.

### Claim 15 (depends on 11)

The method of claim 11, wherein:
the food product comprises soymilk and one or more of: a vegetable ingredient, a fruit ingredient, a seed, an herb, a spice, a plant-based protein ingredient, and a yogurt culture; and
the food product has been pasteurized or homogenized.

### Claim 16 (depends on 11)

The method of claim 11, wherein the selected inflammatory markers include iNOS, COX-2, TNF-alpha, and PGE2.

### Claim 17

A beverage comprising:
at least one gram of protein per eight ounce serving;
a healthy bioactivity index greater than 5;
wherein the beverage is designated as anti-inflammatory.

### Claim 18 (depends on 17)

The beverage of claim 17, wherein the beverage is plant-based.

### Claim 19 (depends on 17)

The beverage of claim 17, wherein the beverage comprises soymilk and one or more of: a vegetable ingredient, a fruit ingredient, a plant-based protein ingredient, a seed, an herb, a spice, and a yogurt culture; and
the beverage has been pasteurized or homogenized.

### Claim 20 (depends on 17)

The beverage of claim 17, wherein the healthy bioactivity index corresponds to Index A or Index B.