1) The study investigated supplementing the feed of laying hens with four omega-3 rich microalgae - Phaeodactylum tricornutum, Isochrysis galbana, Nannochloropsis oculata, and Chlorella fusca - to determine their effects on enriching the omega-3 fatty acid, carotenoid, and sterol content of eggs. 2) Supplementation with all four microalgae increased the omega-3 fatty acid levels in egg yolks, with Phaeodactylum and Isochrysis providing the highest DHA enrichment. Nannochloropsis, Phaeodactylum, and Isoch

1. IMPACT OF FEED SUPPLEMENTATION WITH
DIFFERENT OMEGA-3 RICH MICROOALGAE
SPECIES ON ENRICHMENT OF EGGS OF
LAYING HENS
Charlotte Lemahieu, Charlotte Bruneel,
Romina Termote-Verhalle, Koenraad
Muylaert, Johan Buyse, Imogen Foubert

3. Omega-3
α-Linolenic acid (ALA), 18:3 (n-3)
Eicosapenttaenoic acid (EPA),
20:5 (n-3)
Docosahexaenoic acid (DHA),
22:6 (n-3)
Stearidonic acid (SDA), 18:4 (n-3)
Docosapentaenoic acid (DPA),
22:5 (n-3)

4.  Reduction and prevention of several cardiovascular diseases
 Prevention and treatment of several chronic diseases
 Maternal intake of n-3 LC-PUFA lowers the risk of preterm
birth and low birth weight
 DHA is critical to foetal growth and development of visual
and cognitive functions in foetus and children
Daily intake of n-3 LC-PUFA from 140 to 667 mg/day
 But!!!
 A few countries reach the intake of 250 mg n-3 LC-PUFA per
day
=> Enrich food products with n-3 LC-PUFA has emerged
Why do we supplement Omega-3 Fatty
Acids in egg?

5.  Flaxseed gives mainly rise to enrichment of ALA in the egg
yolk (Fraeye et al., 2012)
 quite inefficient conversion of ALA to EPA and DHA in laying
hens
 Fish oil provides an enrichment of EPA and especially DHA
in the egg yolk (Van Elswyk, 1997)
 The declining fish stocks, the presence of pollutants (heavy
metals and PCBs) in fish
 ≥1.5% fish oil => The off-flavours detected in eggs
Alternative sources of n-3 LC-PUFA must be found
Related researchers

6.  Nannochloropsis was supplemented to laying hens’ feed to
raise the level of n-3 LC-PUFA in the egg yolk
 Fatty acid profile change considerably as well as redness of the
egg yolk increased (Bruneel et al., 2013; Fraeye et al., 2012; Fredriksson et al.,
2006; Nitsan et al., 1999)
 Chlorella and Porphyridium, gave rise to a transfer of
carotenoids from the microalgae to the egg yolk (Ginzberg et al.,
2000; Gouveia et al., 1996)
 Phaeodactylum, as the most promising EPA source and
Isochrysis, as the most promising DHA source, were
recommended (Bruneel et al., 2013)
About microalgal researchers

7.  Investigate the influence of using four different autotrophic
microalgae for n-3 LC-PUFA enrichment in eggs of laying hens.
 Phaeodactylum tricornutum, Isochrysis galbana, Nannochloropsis
oculata and Chlorella fusca
 The four autotrophic microalgae were supplemented in two
different concentrations to the laying hens’ feed
 The fatty acid profile, the carotenoid and sterol content, and
the colour of egg yolk were measured
Objective

9.  Commercial products:
 Phaeodactylum tricornutum (freeze dried biomass, SBAE,
Belgium)
 Nannochloropsis oculata (freeze dried biomass, SBAE, Belgium)
 Isochrysis galbana (freeze dried biomass, Algaenergy, Spain)
 Chlorella fusca (drum dried biomass, Ingrepro, The Netherlands)
 The proximate composition (total lipids, n-3 PUFA,
carotenoids and sterols) and the moisture content was
determined
2.1. Microalgal biomass

10.  72 ISA Brown laying hens
 29 weeks of age
 two hens/cage
 temperature 200C
 16 h of light per day
 ad libitum
 The experimental trial
lasted 42 days
 14 days of adaptation
 28 days of supplementation
with microalgae
2.2. Animals, housing and diets

11. 2.2. Animals, housing and diets (cont.)
No. Treatments Diets
1 Control Without supplement
2 Phaeodactylum triconutum 0,125%
3 Phaeodactylum triconutum 0,25%
4 Nannochloropsis oculata 0,125%
5 Nannochloropsis oculata 0,25%
6 Isochrysis galbana 0,125%
7 Isochrysis galbana 0,25%
8 Chlorella fusca 0,125%
9 Chlorella fusca 0,25%
 After the adaptation period, the hens were randomly assigned to
one of the nine treatment diets (n=8 hens per treatment diet)

12.  Feed intake
 Egg production
 Mortality and morbidity
 Body weight (day 0 and day 28)
2.3. Zootechnical performance of laying hens
were registered on
daily basis during the
entire trial of 42 days

13.  Eggs were collected daily and stored at -200C until further
analysis. The eggs at the end of the supplementation period
were analysed/ or determined for:
 Egg quality parameters
 Total lipid content
 The content and composition of n-3 PUFA, carotenoids and sterols
 To calculate the enrichment of n-3 LC-PUFA in eggs by dietary
supplementation of microalgae, the fatty acid profile of the eggs
at the start (day 0) of the supplementation period was also
analysed.
2.4. Sample collection and storage of eggs

14.  Results were statistically evaluated by two-way analysis
of variance (ANOVA) and a post hoc Tukey’s test with α=
0.05 (Sigmaplot 11, Systat Software, Inc., Illinois, USA).
2.5. Statistical analysis

16. Table 1. Proximate composition of four microalgae: the lipid content
(g/100 g DM), n-3 PUFA content (g/100 g biomass), carotenoid
content (µg/gDM) and sterol content (mg/g DM)
3.1. Proximate composition of n-3 PUFA
rich autotrophic microalgae

17.  The average daily feed intake, egg production, mortality and
morbidity were not significantly affected by supplementation
of microalgae to the laying hens.
 The body weight of the laying hens remained constant, except
for hens supplemented with Nannochloropsis (in both doses),
which displayed a slight decrease (<5%) in body weight.
3.2. Zootechnical performance of laying hens

18. Table 2. Egg quality parameters at the end of microalgal
supplementation period: egg weight, shell weight, albumen weight,
yolk weight (g), shell thickness (mm) and shell strength (N/m)
3.3. Egg parameters

19. Fig.1. Level of omega-3 fatty
acids (mg/egg) in eggs
measured at the end of the
supplementation period
with four microalgae
supplemented in 2 doses
(0.125% and 0.25%); control
(0%)
3.4. Enrichment
of egg yolk
3.4.1. n-3 PUFA
enrichment

20. Table 3. The supplemented n-3 PUFA (mg/100 g feed), the enrichment of
n-3 LC-PUFA (mg/egg) and the conversion efficiency of n-3 LC-PUFA (%)
for the four microalgae supplemented in 2 doses (0.125% and 0.25%)
3.4. Enrichment of egg yolk (cont.)
3.4.1. n-3 PUFA enrichment

21. Table 4. Colour values (Roche and CIELAB-value) of the egg yolk
measured at the end of the supplementation period with four
microalgae supplemented in 2 doses (0.125% and 0.25%)
3.4. Enrichment of egg yolk (cont.)
3.4.2. Yolk colour and carotenoid enrichment
L*: lightness
a*: red-
greeness
b*: yellow-
blueness

22.  As microalgae contain significant levels of health beneficial
phytosterols (Table 1)
 may be transferred to the egg yolk in a similar way as n-3 LC-
PUFA and carotenoids
 may decrease the amount of cholesterol in the egg yolk, giving
the cholesterol lowering properties of phytosterols
 No phytosterols were transferred from the microalgae to the
egg yolk
 Cholesterol was detected in the egg yolk
=> Reducing the cholesterol content in egg yolk is rather
impossible
3.4. Enrichment of egg yolk (cont.)
3.4.3. Yolk sterol enrichment

24.  All four microalgae gave rise to increased levels of n-3 LC-
PUFA in the egg yolk
 The highest enrichment of n-3 LC-PUFA, DHA, was achieved
by supplementation of Phaeodactylum and Isochrysis; a lower
enrichment, Nannochloropsis; and Chlorella gave rise to the
lowest enrichment
 Increase of the carotenoid content was detected by
supplementation of Phaeodactylum, Isochrysis and
Nannochloropsis
 The sterol content of the egg yolk was hardly influenced
Omega-3 rich autotrophic microalgae offer an alternative to
current sources of n-3 LC-PUFA to increase the level of these
fatty acids in the eggs.
The carotenoid enrichment should be taken into account
since the colour shift of the egg yolk could decrease the
consumers’ acceptability.