Lactylates for consistent performance in challenging conditions

CORBION.COM/ALOAPUR

ALOAPUR

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ALOAPUR - today

ALOAPUR powder formulation is suitable for implementation in premix and compound feed:

• Active lactylates

• Carrier powder

• 12 months stable in packaging

• 3 months stable in feed

The active lactylates are made from lactic acid and selected fatty acids (MCFA). They are:

• Fully bio based, patented by Corbion

• Existing products in food and cosmetics

The carrier powder is of diatomaceous earth material

• EU approved as technical Feed Additive

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Aloapur is:

• Fully bio based

• Patented by Corbion

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Aloapur

O

OH

O

OHHO

Fatty acid

+

Lactic acid

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• Blend of C12 & C14 Lactylates ** Only C14 Lactylates

Minimal Inhibitory Concentrations (MIC) are specific to a particular strain and a particular compound, and can depend on: • Current medium composition • Temperature • pH • oxygenation • previous culturing conditions (adaptation)

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Lactylates specifically inhibit Clostridia among other Gram positive bacteria

MIC of C.perfringens

MIC of S. suis MIC of Lactobacillus

spp

Pure C12 lactylate 14 ppm 15 ppm 200 ppm

Pure C14 lactylate

2 ppm 10 ppm 700 ppm

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As individual molecules, lauryl- (C12) and myristoyl (C14) Lactylates inhibit various Clostridia species at low concentrations (<10 ppm) in vitro:

The bacteriostatic effect is fully maintained in the formulation of ALOAPUR lactylates

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Lactylates specifically inhibit Clostridia among other Gram positive bacteria

Growth curves of C. perfringens with increasing levels of lactylates

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Lactylates have superior bactericidal activity against of Clostridium

• Free Fatty Acids (MCFA) below C12 can reduce bacteria at low pH

• Converting an MCFA into a lactylate enhances this activity

• Converting an MCFA into a lactylate makes is less pH dependent

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Lactylates are stronger and less pH-dependent anticlostridium molecules than fatty acids

0

1

2

3

4

5

6

7

C12 Lac(1) C14 Lac(1) C12 FA C14 FA

Log

Re

du

ctio

n C

FU/m

l

Reduction of C. perfringens 10 min, pH 6,5

100 ppm

50 ppm

0

1

2

3

4

5

6

7

C12 Lac(1) C14 Lac(1) C12 FA C14 FA

Log

Re

du

ctio

n C

FU/m

l

Reduction of C. perfringens 10 min, pH 4.5

100 ppm

50 ppm

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Benchmarking available feed products: Lactylate products effect a stronger kill on Clostridia than free fatty acid products

0

1

2

3

4

5

6

C12 FFA (competitor 1) 100 ppm

C8-C12 FFA (competitor 2) 100

ppm

C8-C18:1 FFA (competitor 2) 100

ppm

Mix product (competitor 3) 100

ppm

Current Aloapur active 100 ppm

Next Gen Aloapur active 100 ppm

Log

Re

du

ctio

n (

CFU

/ml)

Kill Clostridium perfringens at pH 6.5

10 min

60 min

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The mechanism of antibacterial activity was visualized with microscopic methods as well:

• C. perfringens was grown to high density in vitro

• Green dye passes intact cell membranes

• Red dye passes disrupted cell membranes

Conclusions:

• In vitro, lactylates reduce C. perfringens

• The mechanism likely involves disruption of the cell membrane

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Lactylates disrupt bacterial cell membranes

0 ppm lactylates

500 ppm lactylates

50 ppm lactylates

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The overall effect of lactylates on gut flora was characterized in vitro:

• Cecal microflora of 18 day old broilers was maintained in vitro

• This complex microbial culture was exposed to lactylates and various antibiotics

Conclusions:

• Lactylates share with antibiotics a general effect on the microflora (shift type 1)

• Lactylates induce a unique type of shift not seen in any of the tested antibiotics (shift type 2)

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Lactylates exert a pressure on gut flora like antibiotics do, in a distinctive pattern

Lactylates Antibiotics

Controls

Gu

t fl

ora

sh

ift

typ

e 1

Gut flora shift type 2

In vitro microflora: composition analysis by I-chip, Principal Component Analysis, visualizing the two most prominent type of composition shifts

*References: TNO website, TNO report

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The overall effect of lactylates on gut flora was characterized in vitro:

• Cecal microflora of 18 day old broilers was maintained in vitro

• This complex microbial culture was exposed to lactylates and various antibiotics

Conclusions:

• Lactylates share with antibiotics a general effect on the microflora (shift type 1)

• Lactylates induce a unique type of shift not seen in any of the tested antibiotics (shift type 2)

• Lactylates specifically reduce C. perfringens relative to overall flora

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Lactylates specifically suppress the numbers of C.perfringens in the context of in vitro gut flora

In vitro microflora: QPCR quantification of C.perfringens and overall bacterial numbers

1E+00

1E+01

1E+02

1E+03

1E+04

1E+05

1E+06

1E+07

1E+08

C. perfringens all bacteria

bct

eria

nu

mb

ers

(CFU

)

Control culture

ALOAPUR lactylates

*References: TNO website, TNO report

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The mechanism(s) of the direct antibacterial effect were mapped and compared to antibiotics:

• C. perfringens was grown in vitro and exposed to Lactylates and various antibiotics

• The cellular responses were mapped through gene expression profiling

Conclusions:

• Lactylates provoke a different cellular response compared to any of the tested antibiotics

• Based on this finding and their molecular structure, lactyates will not contribute to antibiotic (cross-) resistance

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Lactylates do not contribute to antibiotic (cross-) resistance

Ce

llula

r re

spo

nse

typ

e 1

Cellular response type 2

Lactylates

Antibiotics

Monoculture C.perfringens cellular responses: RNA expression profiling of monoculture cells exposed to antimicrobials, lactylates in dark blue oval, tylosin in red, monensin in light blue, salinomycin in yellow, ampicillin in green

*References: TNO website, TNO report

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0%

5%

10%

15%

20%

25%

Concentration of Lactylates (% of in-feed dosing level)

C12-1 Lactylate C14-1 Lactylate

The reach of lactylates in the broiler digestive tract was established by WUR Wageningen:

• Broilers were fed high viscosity diets, dosed with Aloapur

• Lactylate levels were measured in the feed, in digesta and in the blood

Conclusions:

• Lactylates are present in significant concentration in the GI tract down to the jejunum

• Specific anti-clostridium gut flora shifting effect is applied in upper GI tract, where numbers of clostridia should be low

• Aloapur likely serves to support upper gut microflora as barrier against oral Clostridium

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Lactylates reach the small intestine (1)

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The reach of lactylates in the broiler digestive tract was established with WUR Wageningen:

• Broilers were fed high viscosity diets, dosed with Aloapur

• Lactylates levels were measured in the feed, in digesta and in the blood

Conclusions:

• Lactylates are present in significant concentration in the GI tract down to the jejunum

• Specific anti-clostridium gut flora shifting effect is applied in upper GI tract, where numbers of Clostridia should be low

• Aloapur likely serves to support upper gut microflora as barrier against descending Clostridium

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Lactylates reach the small intestine (2)

0

5

10

15

20

25

30

35

40

45

50

Duodenum Proximal Jejunum Distal Jejunum

Concentration of lactylates (ppm)

C12-1 Lactylate

C14-1 Lactylate

C14-1 Lactylate C. perfringens MIC In vitro*

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Results - Animal Live Trials *

* The results of the Animal Trials were performed with a natural challenge like: - Homogenized litter - Feed e.g. corn/wheat No Clostridia challenge was induced in the next cases.

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The effect of dosing constant levels of ALOAPUR was investigated:

• Ross 308 broilers were reared in floor pens and fed a wheat/corn/soy diet

• ALOAPUR was dosed at 0,25% to 1,5% throughout the rearing period

Conclusions:

• At equal growth rates, ALOAPUR decreased FCR in a linear response, by over 5%

• No harmful effects were observed at super dosing of ALOAPUR

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Aloapur supports high feed efficiency

94%

95%

96%

97%

98%

99%

100%

000% 000% 001% 001% 001% 001% 002%

FCR

as

% o

f N

eg.

Co

ntr

ol

ALOAPUR dosing

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Aloapur supports optimal performance in mildly challenging conditions

The effect of dosing stepwise levels of ALOAPUR was investigated:

• Ross 308 broilers were reared on used litter without disinfection or litter treatment

• ALOAPUR was dosed at: 0,5% to day 21 – 0,2% to day 35 “Stepwise” 0,5% to day 21 – 0,1% to day 35 “Stepwise” 0,2% throughout day 35 rearing period “Constant” 0,2% to day 21 – 0,1% to day 35 “Stepwise”

Conclusions:

• Aloapur dosing resulted in 3.4% to 4.9% increase in BWG and improved the FCR with -1.2% to -4.4%

• Stepwise decreasing dose important to achieve overall ROI by ALOAPUR

-06

-04

-02

00

02

04

06

BWG FCR

Pe

rfo

rman

ce d

iffe

ren

ce t

o c

on

tro

l (%

)

Period 1-35 days

Aloapur 5.0 - 5.0 - 2.0

Aloapur 5.0 - 5.0 - 1.0

Aloapur 2.0 - 2.0 - 2.0

Aloapur 2.0 - 2.0 - 1.0

*

*

*

*

*

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The effect of dosing stepwise levels of ALOAPUR was investigated:

• Ross 708 broilers were reared in sub-optimal conditions

• ALOAPUR was dosed at 2kg/Ton and compared to:

• Antibiotics

• Essential oil/MCFA mixtures

Conclusions:

• 2 kg/Ton dosing resulted in significant performance improvement

• Aloapur outperformed competing alternatives at recommended dosing levels

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Aloapur supports optimal performance in challenging conditions at 2kg/Ton

*Reference: World Poultry article by H Van Iersel en K Wijma (2014)

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Aloapur in piglets

Improvements in Aloapur group

End weight +3.4 %

FCR -2.86 %

The effect of dosing Softacid Piglet together with Aloapur: • Piglets were reared in industrial

conditions

• ALOAPUR was dosed at 2kg/Ton and compared to a commercial blend of organic acids and MCFA

Conclusions: • 2 kg/Ton dosing resulted in significant

performance improvement • Aloapur outperformed competing

alternative at recommended dosing levels

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Aloapur in piglets

The effect of dosing Aloapur compared to dosing antibiotics • Aloapur dosed at 2 kg/Ton was evaluated commercial alternative in industrial

conditions (2 stables next to each other)

• During the trial: outbreak of viral infection PRRS in control group which resulted in: Reduced growth

High mortality rates Increased respiratory problems

• No outbreak in Aloapur control group

Conclusions: • Aloapur improves health by reducing gram+ infection pressure which makes the

piglets more resistent to other infections

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Farm had problems with Ileitis and Clostridia. 925 fattening pigs were divided into 2 groups with each 6 pens containing 13 - 15 pigs

Treatment: • Control • Aloapur 2 kg/MT

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Aloapur improves the overall Health of Fattening Pigs Private Farm in Spain - 925 Fattening pigs

0

0,2

0,4

0,6

0,8

1

1,2

1,4

1,6

Control Aloapur

Mo

rtal

ity

(%)

Mortality per treatment

Clostridia

Ileitis

Conclusion Aloapur controls gut bacteria which causes intestinal problems and economic losses. Aloapur reduces the mortality rate and therefore increases the amount of animals to the slaughter house.

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Comparative field trial in piglets, Belgium 2017

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Commercial acid blend

Brenntag Concept

(NS Piglet 4 kg/mT + Aloapur 2 kg/mT)

D

Improvement

(%)

Weight gain (g/d) 581a 628b +8,1

Feed intake (g/d) 879a 939b +6,8

Feed conversion 1.514 1.500 -0,9

AB use 5 days AMX No AB needed

Piglets, day 21 – 69, 576 piglets

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Comparative field trial in piglets, Belgium 2017

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Standard diet: control

Brenntag Concept

(NS Piglet 4 kg/mT + Aloapur 2 kg/mT)

D

Improvement

(%)

Start weight (kg) 6.46 6.45

End weight (kg) 14.86b 16.58a 11.6

Daily weight gain (g) 255b 307a 20.4

Daily feed intake (g) 332b 367a 10.5

FCR 1.30b 1.20a - 7.7

AB use 4,7 days AMX No AB use

576 piglets, weaned at 21 days, monitored 54 days Control feed: traditional acidifier diet, stable switch compared to previous trial to eliminate stable effect

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Comparative field trial in piglets, Belgium 2017

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Standard diet Brenntag Concept

(NS Piglet 4 kg/mT + Aloapur 2 kg/mT)

D

Improvement

(%)

Initial weight (kg) 5.04 5.05

Final weight (kg) 7.19 7.43 3.3

Daily gain (g) 254 270 6,3

Feed conversion 1.25 1.22 -2.4

% Mortality 2.45 2.09 -14.7

28-42 days after weaning

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Brenntag concept tested in sows

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Neubacid Soft piglet (4 kg/mT) + Aloapur (2 kg/mT) before and during lactation:

2 treatments: standard control group and Brenntag concept group, weaned at 21 days, 500 sows

Results in sows:

Weight drop during lactation reduced with 5 kg

11% more sows without deathborn piglets

24% more sows with less than 2 death born piglets

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Brenntag concept tested in sows & piglets

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Neubacid Soft piglet (4 kg/mT) + Aloapur (2 kg/mT) during before and during lactation:

2 treatments: standard control group and Brenntag concept group, weaned at 21 days, 500 sows

Results in piglets:

Weaning weight: +0,35 kg/piglet

Less mortality in piglets (- 4,22%)

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Aloapur in Rabbits

Coccidiose is major disease in rabbits resulting in: - Poor growth - High FCR - High mortality => significant losses

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Aloapur in Rabbits: test 1

432 Cross-bred New ZealandxCalifornian rabbits 3 groups (18 cages consisting out of 8 animals) Well ventilated and clean cages 3 Treatments: 1. T1: control group: 120 ppm colistin 100 ppm tiamulin 100 ppm Zinc bacitracin 2. T2: negative control 3. T3: Aloapur group: 2 kg/mT Aloapur 80 ppm colistin

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Aloapur in Rabbits: test 1

Medicated diet No medicated diet Aloapur diet

Bodyweight d29(kg)

1.979 1.971 1.975

Feed intake (kg/day)

3.273a 3.311a 3.096b

FCR (d0-29) 3.3a 3.3a 3.1b

Mortality (%) 4.8a 16.3b 4.8a

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Aloapur in Rabbits: test 2

198 Cross-bred New ZealandxCalifornian rabbits 3 groups (11 cages consisting out of 6 animals) Sub clinical conditions and ‘dirty ‘ cages 3 Treatments: 1. T1: control group: 120 ppm colistin 100 ppm tiamulin 100 ppm Zinc bacitracin 2. T2: negative control 3. T3: Aloapur group: 2 kg/mT Aloapur

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Aloapur in Rabbits: test 2

Medicated diet No medicated diet Aloapur diet

Bodyweight d29(kg)

1.97a 1.71b 1.90ab

Feed intake (kg/day)

3.06a 3.17b 3.03a

FCR (d0-29) 2.80a 3.10b 2.9a

Mortality (%) 1.5a 30.3b 6.1a

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ALOAPUR

Chemistry

New molecules in animal nutrition

Safe, food grade product

EU Feed Material Catalogue 11.4.7

Product

Lactylates on carrier

Free flowing powder

EU complementary feed

Functionalities

Modulates gut flora

Robust guts for challenging conditions

Consistently high poultry performance