Repox50

A novel, natural polyphenol partial replacement for vitamin E in feeding

diets

Presentation overview The problem

– Oxidative stress

The solution

– Antioxidants

Our recommended solution

– Repox50

Repox50

The problem

– Oxidative stress

The solution

– Antioxidants

Our recommended solution

– Repox50

Repox50

Reactive oxygen species Reactive oxygen species (ROS) are chemically-reactive molecules

containing oxygen. Examples include oxygen ions and peroxides.

Increased ROS can result in significant damage to cell structures. This cumulates into a situation known as oxidative stress.

Antioxidants are used to counteract the effect of these increased ROS levels often in combination with the addition of extra minerals and vitamins to the diet.

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Some causes of oxidative stress?

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Problems resulting from oxidative stress

Growth inhibition

Muscle degeneration, leading to inferior meat quality

Suppression of the immune system response

Loss of flavour, texture, appearance and in some cases nutritional value of the meat

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The problem

– Oxidative stress

The solution

– Antioxidants

Our recommended solution

– Repox50

Repox50

What are antioxidants? Antioxidants are phytochemicals, vitamins and other nutrients

that protect our cells from damage caused by free radicals.

In vitro en in vivo studies have shown that antioxidants help prevent the free radical damage that is associated with disease.

Antioxidants have the ability to neutralize free radicals without becoming a free radicals themselves, however, when the antioxidant neutralises a free radical it becomes inactive. Therefore we need a continuous supply of antioxidants.

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Antioxidants in action

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Some commonly used antioxidants Vitamin E

– A fat-soluble antioxidant that stops the production of reactive oxygen species formed when fat undergoes oxidation.

Vitamin C

– Vitamin C or L-ascorbic acid or L-ascorbate is an essential nutrient for humans and certain other animal species, in which it functions as a vitamin.

Selenium

– Selenium is necessary for cellular function in most, if not all, animals, forming the active centre of the enzymes glutathione peroxidase and thioredoxin reductase.

Repox50

The problem

– Oxidative stress

The solution

– Antioxidants

Our recommended solution

– Repox50

Repox50

Repox50 the natural choice Super concentrated polyphenols extracted from specifically

selected botanical sources.

Works in synergy with Vitamin E, able to replace up to 50% resulting the same or superior protective effect, thus creating more cost effective inclusion into in poultry diets.

Sustainably produced from natural renewable resources.

Safe and easy to handle.

Repox50

Repox50 Heat StabilityOVERVIEWTwo representative samples of Repox50 were received from

Phytologica Ltd, batch number LAB/PHY01. This sample was separated into 2 samples for test purposes. Sample 1 and Sample 2.

• Sample 1 was subjected to an increase in temperature for 60 minutes in an autoclave at a temperature of 120°C.

• Sample 2 was untreated.• Sample 1 and Sample 2 were analysed for polyphenols using

a cyclic voltammetry technique (CV) used routinely in the Citrox QC laboratory.

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Repox50 Heat StabilityLAB/PHY01 - Sample 1 (120°C)

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MondaydEC1514W.efwdat : Page 21

Mode: Cyclic, Initial E : -1200mV, Final E : -1200mV, R ate: 80mV/ s, S tep W : 25ms, Upper E : 1200mV, Lower E : -1200mV, Cycles: 5 (Mon, Dec 15, 2008, 4:00:59 P M)

Some data points in the recording are out of range. T his may affect the results.

-40

-20

0

20

40

i 1 (

A)

-1.2 -0.8 -0.4 0 0.4 0.8 1.2E (V)

Repox50 Heat StabilityLAB/PHY02 - Sample 2 (untreated)

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MondaydEC1514WJE.efwdat : Page 26

Mode: Cyclic, Initial E : -1200mV, Final E : -1200mV, R ate: 80mV/ s, S tep W : 25ms, Upper E : 1200mV, Lower E : -1200mV, Cycles: 5 (Mon, Dec 15, 2008, 4:08:13 P M)

Some data points in the recording are out of range. T his may affect the results.

-40

-20

0

20

40

i 1 (

A)

-1.2 -0.8 -0.4 0 0.4 0.8 1.2E (V)

Repox50 Heat StabilityConclusion

• “there were no discernable differences in the voltammograms which can be thought of as “fingerprints” of the polyphenol activity”

Testing carried out by Dr. David J. Lax BSc MRSC PhD Chemical Research Services

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Some scientific evidence Sparing effect of procyanidins from Vitis vinifera on vitamin E: in vitro

studies

– Carini M, Maffei Facino R, Aldini G, CalloniI MT, Bombardelli E, Morazzoni P, 1998 : The protection of polyunsaturated fatty acids in micellar systems against UVB-induced photo-oxidation by procyanidins from Vitis vinifera L., and the protective synergy with vitamin E. International J. of Cosmetic Science, Volume 20 Issue 4 Page 203-215,August 1998.

Select flavonoids and whole juice from purple grapes inhibit platelet function and enhance nitric oxide release

– Freedman JE, Parker C 3rd, Li L, Perlman JA, Frei B, Ivanov V, Deak LR, Iafrati MD, Folts JD. , 2001. Select flavonoids and whole juice from purple grapes inhibit platelet function and enhance nitric oxide release. Circulation. 2001 Jun 12;103(23):2792-8.

Catechins and procyanidins in Mediterranean diets– AUGER Cyril, AL-AWWADI Najim, BORNET Aurélie, ROUANET Jean-Max, GASC Francis, CROS Gerard ,

TEISSEDRE Pierre-Louis : Catechins and procyanidins in Mediterranean diets Food research International 2004, vol. 37, no3, pages. 233-245 [13 page(s) (article)] (1 p.1/4)

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In vitro antioxidant trialsTrial ObjectiveTo measure the antioxidant activity at 0%, 25%, and 50% of the vitamin E substitued by

Repox50 and compare with control. Growth Control: No antioxidant added 100% Vitamin E: 0% substitution 75% Vitamin E: 25% substition 50% Vitamin E: 50% substitution

Conclusion 100% Vitamin E antioxidant activity

is 250% higher than negative control Replacement of 25% and 50% of

Vitamin E by Repox50 the antioxidant activity is still identical to 100% Vitamin E

Increase of antioxidant activity compared to no antioxidant addition

34.00%

33.00%

33.00%

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Poultry Applications34.00%

33.00%

33.00%

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Poultry diet trialsEffect of Repox50 and Vitamin E on Growth Performance, Nutrient

Digestibility in Chickens 120 one-day-old male broiler chicks  were obtained from a commercial hatchery and

housed in an environmentally controlled room over a period of 3 weeks. All diets were formulated to meet or exceed the minimum NRC (1994) requirements for broiler chickens.

At the end of the experimental period, birds were weighed, and feed consumption was recorded

The addition of increasing concentration of Repox50 in the chicken diets did not impair growth performance (BW, feed consumption, and feed efficiency) compared with the birds fed the non supplemented and supplemented vitamin E diets

Antioxidant activity in vitamin E and Repox50 diets exhibited significantly higher scavenging free radical capacity than controls.

Similarly, the birds fed vitamin E and Repox50 diets exhibited significantly higher scavenging free radical capacity in excreta than those fed control diets.

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Evaluation of Repox50 in Broilers under research station circumstances

Trial No. AB11238B

BANGKOK ANIMAL RESEARCH CENTER CO., LTD. HEAD OFFICE

487/1 SI AYUTTHAYA ROAD, KHWAENG THANON PHAYATHAI, KHET RATCHATHEWI,

BANGKOK 10400, THAILAND

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Trial DetailObjectiveThis trial was conducted to evaluate the effect of replacing vitamin E by

Repox 50 on performance of broilers. Materials and methodsThe trial was conducted at BARC research facility during July 6 – August 10,

2011. Six hundred newly hatched male broiler chicks of commercial strain (AA Plus) were randomly allocated to 3 treatments with 8 replications in a randomized complete block design experiment, using 25 chicks per experimental unit. Practical corn-soybean diet with 100 g/t added vitamin E 50 was used as control diet and two levels of replacing of vitamin E 50 by Repox50. Treatment diets were presented as below.

Diet 1 = Control diet: practical diet with 100 g/tone added Vitamin E 50Diet 2 = Replacing 25% of added Vitamin E of diet 1 by Repox 50Diet 3 = Replacing 50% of added Vitamin E of diet 1 by Repox 50

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Composition of starter and grower dietsCode Ingredient

Price   Starter Grower

(Baht / kg)   0-18 d 18-35 d    Control diet Control diet

B103-1 Corn 7.35% 11.50   49.09 57.67

B422 Soybean meal (DH) 49.0% 14.07   31.56 25.94

B417-1 Full fat soybean 36.0% 19.17   10.00 8.00

B202 Soybean oil 40.00   4.82 4.67

B601-2 MCP 18.50   1.78 1.44

B602-2 Lime stone 39% Ca 1.40   1.06 0.90

B603 Salt 4.00   0.33 0.23

B908 Pellet binder 70.00   0.30 0.30

B703 Vitamin/mineral permix (100g/t of vitamin E 50) 55.00   0.20 0.20

B502 DL-Methionine 150.00   0.27 0.18

B501 L-Lysine HCl 72.00   0.16 0.07

B503 L-Threonine 78.00   0.03 -

B605 Sodium bicarbonate 15.00   0.20 0.20

B606 Choline chloride 60% 64.00   0.10 0.10

B804 Toxin binder 200.00   0.05 0.05

B910 Coccidiostat (Salinomycin 12%) 250.00   0.05 0.05

 TOTAL   100.00 100.00

  COST Baht / kg   15.47   14.93

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Nutrients of starter and grower diets

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NUTRIENT Unit  

1 Weight Kg   1 1

2 Dry matter %   87.85 87.623 ME. For poultry Kcal/kg   3050 3100

9 Crude protein Kcal/kg   23.0 20.011 Crude fat Kcal/kg   9.05 8.73

12 Linoleic acid %   4.29 4.16

13 Crude fiber %   3.22 3.0117 Dig.Lys (Poultry) %   1.24 1.00

18 Dig.Met (Poultry) %   0.58 0.4619 Dig.Cys (Poultry) %   0.31 0.29

20 Dig.M+C (Poultry) %   0.89 0.75

21 Dig.Thr (Poultry) %   0.77 0.6522 Dig.Trp (Poultry) %   0.20 0.17

23 Dig.Arg (Poultry) %   1.42 1.22

24 Dig.Val (Poultry) %   0.96 0.8425 Dig.Ile (Poultry) %   0.83 0.72

26 Dig.Leu (Poultry) %   1.69 1.5546 Lysine %   1.41 1.14

47 Arginine %   1.59 1.36

48 Methionine %   0.62 0.5049 Met + Cys %   1.00 0.84

50 Cystine %   0.38 0.35

52 Threonine %   0.92 0.78

53 Tryptophan %   0.27 0.23

55 Histidine %   0.62 0.5556 Isoleucine %   1.02 0.88

57 Leucine %   1.89 1.71

58 Valine %   1.11 0.97

59 Phenylalanin %   1.07 0.9466 Calcium %   0.86 0.72

67 Phosphorus-total %   0.75 0.66

68 Phosphorus-avail %   0.43 0.36

69 Non phytate %   0.50 0.4278 Sodium %   0.20 0.1679 Chloride %   0.23 0.18

80 Salt %   0.36 0.27             

Statistical AnalysisFeed intake, body weight gain, feed conversion ratio, liability, FRAP value

and TBA value were determined and were subjected to ANOVA as RCBD. Significant differences among treatment groups were detected by LSD tests (least square deviation).

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ResultsTable1 Evaluation of Repox 50 in broilers1 under research station

circumstances (0-18 days of age).

1 Male broilers (Arbor Acres Plus).2 Feed conversion ratio corrected for mortality and culls.3 Feed conversion ratio uncorrected for mortality and culls.

Treatment Initial Final Body Feed Feed Feed Livability

Diet Repox 50 body weight weight intake conversion conversion

Group With Vit E 50 (replacing of weight gain gain ratio2 ratio3

100% Vit E 50) (g) (g) (g) (g) (%)

1 Control diet - 44 838 793 957 1.207 1.210 99.5

2 As diet 1 25 % 44 835 790 956 1.210 1.217 99.0

3 As diet 1 50 % 44 833 789 964 1.222 1.228 99.0

P-value 0.7026 0.6850 0.4195 0.1181 0.1213 0.8815

Pooled SEM 3.576 3.564 4.448 0.005 0.006 0.809

C.V.% 1.21 1.27 1.31 1.15 1.33 2.31

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Table 2 Evaluation of Repox 50 in broilers1 under research station circumstances (18-35 days of age).

1 Male broilers (Arbor Acres Plus).2 Feed conversion ratio corrected for mortality and culls.3 Feed conversion ratio uncorrected for mortality and culls.

Treatment Initial Final Body Feed Feed Feed Livability

Diet Repox 50 body weight weight intake conversion conversion

Group With Vit E 50 (replacing of weight gain gain ratio2 ratio3

100% Vit E 50) (g) (g) (g) (g) (%)

1 Control diet - 44 838 793 957 1.207 1.210 99.5

2 As diet 1 25 % 44 835 790 956 1.210 1.217 99.0

3 As diet 1 50 % 44 833 789 964 1.222 1.228 99.0

P-value 0.7026 0.6850 0.4195 0.1181 0.1213 0.8815

Pooled SEM 3.576 3.564 4.448 0.005 0.006 0.809

C.V.% 1.21 1.27 1.31 1.15 1.33 2.31

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Table 3 Evaluation of Repox 50 in broilers1 under research station circumstances (0-35 days of age).

1 Male broilers (Arbor Acres Plus).2 Feed conversion ratio corrected for mortality and culls.3 Feed conversion ratio uncorrected for mortality and culls.

Treatment Initial Final Body Feed Feed Feed Livability

Diet Repox 50 body weight weight intake conversion conversion

Group With Vit E 50 (replacing of weight gain gain ratio2 ratio3

100% Vit E 50) (g) (g) (g) (g) (%)

1 Control diet - 44 2565 2521 3798 1.507 1.524 98.0

2 As diet 1 25 % 44 2571 2527 3826 1.515 1.521 98.5

3 As diet 1 50 % 44 2580 2535 3831 1.511 1.526 98.0

P-value 0.7960 0.7926 0.4966 0.5555 0.9726 0.9641

Pooled SEM 15.253 15.243 20.676 0.005 0.015 1.508

C.V.% 1.68 1.71 1.53 0.91 2.70 4.34

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Table 4 Evaluation of Repox 50 on fat oxidation ability in broilers1 under research station circumstances.

1 Male broilers (Arbor Acres Plus).2 Thiobarbituric acid3 Ferric reducing antioxidant power

Treatment Initial Final Body Feed Feed Feed Livability

Diet Repox 50 body weight weight intake conversion conversion

Group With Vit E 50 (replacing of weight gain gain ratio2 ratio3

100% Vit E 50) (g) (g) (g) (g) (%)

1 Control diet - 44 2565 2521 3798 1.507 1.524 98.0

2 As diet 1 25 % 44 2571 2527 3826 1.515 1.521 98.5

3 As diet 1 50 % 44 2580 2535 3831 1.511 1.526 98.0

P-value 0.7960 0.7926 0.4966 0.5555 0.9726 0.9641

Pooled SEM 15.253 15.243 20.676 0.005 0.015 1.508

C.V.% 1.68 1.71 1.53 0.91 2.70 4.34

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Fish Applications

Repox50 will enhance colour and shelf life via replacing a portion of Vitamin E within the diet

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Polyphenols can improve body color and meat brilliance of cultured fishes, preventing decrease in astaxanthin levels in the fish.

Polyphenols added by weight from 0.004% 40ppm to 0.04% 400ppm.

Sakiura US 6,509,030, B2

Cultured fish carotenoid and polyphenol added feed for improving fish body color tone and fish

meat brilliance  

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Repox50

Meat Applications

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Some scientific evidence Ahn, J., Grün, I.U., Fernando, L.N. 2002. Antioxidant properties of natural plant

extracts containing polyphenolic compounds in cooked ground beef. J. Food Sci. 67(4), 1364-1369.

Ahn, J., Grün, I.U., Mustapha, A. 2007. Effects of plant extracts on microbial growth, color change, and lipid oxidation in cooked beef. Food Micro. 24, 7-14

Bekhit, A.E.D., Geesink, G.H., Ilian, M.A., Morton, J.D., Bickerstaffe, R. 2003. The effects of natural antioxidants on oxidative processes and metmyoglobin reducing activity in beef patties. Food Chem. 81(2), 175-187

Brannan, R.G., Mah, E. 2007. Grapeseed extract inhibits lipid oxidation in muscle from different species during refrigerated and frozen storage and oxidation catalyzed by peroxynitrite and iron/ascorbate in a pyrogallol red meat model system. Meat Science. 77(4), 540-546.

Jayaprakasha, G.K., Singh, R.P., Sakariah, K.K. 2001. Antioxidant activity of grape seed (vitis vinifera) extracts on peroxidation models in vitro. Food Chem. 73(3), 285-290.

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Factors affecting meat colour• Vitamin E feeding of cattle

– Prevents oxidation; retards conversion of myoglobin to metmyoglobin

• Bacteria– Produce metmyoglobin, choleglobin, and sulfmyoglobin

pigments• Curing

– Nitrosylhemochromogen is the stable cured meat pigment

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Repox50 improves meat colour

Factors affecting meat colourExposure to long-term or short term stress• Effects glycogen content of muscle and ultimate pH of muscle• Long Term Stress: DFD (dark cutter)Transport, Hunger, Fear,

Aggression• Ultimate pH above 5.9 (beef), 6.5 (pork)• Short Term Stress: PSE Usually only problematic in pork • Ultimate pH below 5, generally problem can be overcome with

enhancement

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Thank you for your time

Any questions?

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