FEED TECHNOLOGY
Shrimp pigmentation with
natural carotenoids
By Herv Lucien-Brun and Frdric Vidal
Natural alfalfa concentrate in finisher feed fed during the last month of the grow-out cycle significantly
increases the reddish colouration of shrimp after cooking.
Dr Gerard Cuzon and the IFREMER team in Tahiti, French Polynesia
studied the effect of adding a natural source of carotenoids from alfafa
concentrate on shrimp pigmentation. The product is obtained by the
dehydration of this widely used forage. The extraction process uses
water and heat application.
In a series of trials, conducted in clear water tanks with controlled
parameters (36ppt and 26C), the effect of the alfalfa concentrate
Pigmentech (Aqua Techna, France) was tested on marine shrimp Penaeus
stylirostris, at three inclusion rates T (0% Pigmentech), A (4%
Pigmentech) and B (8% Pigmentech). There were 6 replicates.
Variation of the colouration between the three batches after 21 days of experimentation
Shrimp were individually weighed at the start of the experiment
and fed 3-4 times/day at 5% body weight per day. Shrimp were sampled
on day 7 and day 21 and two shrimp were sampled from each tank per
time (6 replicates and thus 12 shrimp per treatment). The analysed
parameters were:
Pigmentation: For this purpose, 6 shrimp from each treatment were
immersed in boiling water (5g/l of salt) for 3 minutes. Subsequently,
the shrimp were placed on a black area and/or a white zone in order
to evaluate colouration using the Roche index: SalmoFan (a graduated
34 | May/June 2006 AQUA Culture AsiaPacific Magazine
Table 1. Changes in colouration after 21 days of experiment
T (0% inclusion)*
26.4 1.3
A (4% inclusion)*
28.3 1.1
B (8% inclusion)*
30.2 0.9
*Percent inclusion of the alfafa concentrate Pigmentech in diet of shrimp
Figure 1. Effect on the shrimp colouration
pink color chart from 20 to 34, see photo). Colour was evaluated by a
team of 10 people.
Zootechnical: The growth rate, survival and number of molts were
analysed. After 7 days of experiment, there was no statistical difference
between treatments. However, at day 21, the results were as shown in
Table 1 and after 21 days of experiment, colour was significantly
enhanced as seen in cooked shrimp.
The addition of the alfafa concentrate also showed a very positive
effect on animal responses, especially in terms of growth as shown in
Figure 2. This improvement in the growth rate may be explained by the
digestibility of the proteins contained in the alfalfa. However, it could
also be attributed to the high content of natural antioxidants and
Figure 2. Effects on the growth
Shell
T (0%)
A (4%)
B (8%)
P (from pond culture)
Epidermis
T (0%)
A (4%)
B (8%)
P (from pond culture)
Hepatopancreas
T (0%)
A (4%)
B (8%)
P (from pond culture)
Number of shrimp
5
6
6
6
Number of shrimp
5
6
6
6
Number of shrimp
6
6
6
6
Q (g)
0.6 0.1
0.7 0.2
0.8 0.3
0.8 0.4
Q (g)
24.8 2.6
32.8 12.1
42.2 4.8
27.8 9.9
Q (g)
18 10
81 66
104 78
340 90
C (g/g)
18.7 10.8
12.2 2.3
20.8 10.7
13.9 6.3
C (g/g)
510 141
890 221
1,383 318
781 235
C (g/g)
28 15
119 95
156 115
1,144 520
Table 3. Distribution of the various forms of astaxanthin in the
hepatopancreas and in the epidermis of the shrimp sample
Percent of total carotenoids in the hepatopancreas
Treatments
T (0%)
A (4% )
Free astaxanthin
43.9
31.9
Mono-ester astaxanthin
8.7
24.8
Di-ester astaxanthin
12.0
15.0
Total astaxanthin
64.6
71.7
Percent of total carotenoids in the epidermis
Treatments
T (0%)
A (4% )
Free astaxanthin
48.4
55.3
Mono-ester astaxanthin
17.1
18.6
Di-ester astaxanthin
18.4
12.3
Total astaxanthin
83.9
86.2
B (8%)
29.4
24.0
18.0
71.4
B (8%)
48.5
26.4
10.9
85.8
Q is the quatity of carotenoids as g/g in the sample (shell and epidermis, 3.6cm2 per sample and the
total hepatopancreas). C is the concentration of carotenoids as g/g of dry weight of tissue after extraction.
essential fatty acids. Nevertheless, no effect was observed in terms of
molting rate or survival, during this period.
This first study showed that shrimp colouration was enhanced by
the use of the concentrate in the feed, even at low rates of inclusion
(4 and 8%). This showed that shrimp fed with these diets could
accumulate the carotenoid pigments and demonstrate similar beneficial
effects. Similar results were obtained with Penaeus vannamei by
separate teams in Thailand and China.
Analysis of carotenoid contents
The purpose of the second work carried out by O. Berticat, R. Castillo
and G. Ngre-Sadargues from the Laboratory of Ecophysiology of the
Invertebrate, University of Montpellier, France was to investigate factors
controlling the nature of the pigments involved in pigmentation of
shrimp. The pigments present in the concentrate are mainly yellow
(lutein) and orange (carotene). Thus, the objective was to determine
the nature of the pigments obtained after bioconversion by the shrimp.
During the experiment carried out in Tahiti, six shrimp in each
treatment T, A and B, were sampled and deep frozen at -80C. At the
same time, shrimp (group P) were sampled from semi-intensive earthen
ponds in a farm in Tahiti.
For each individual animal, the total carotenoid inclusion level and
the qualitative analysis of these pigments were carried out by
chromatography on tissues. These results in Table 3 showed an increase
in the carotenoid concentration in the epidermis from group T to B. The
same tendency could be noted in the hepatopancreas.
In each group, the exoskeleton was near transparent with a very
low concentration of carotenoids. The epidermis is the richest tissue
in pigments. This concentration was directly related to the level of
pigments present in feed. The pigment content in the hepatopancreas
showed significant individual variations. It also showed increasing
concentrations with the increases in the level of pigment in food.
In spite of the small number of analyses carried out, the results
indicated that P. stylirostris fed pellets supplemented with carotenoids
absorbed the pigment in the feed, which after bioconversion, accumulated
in the hepatopancreas before being transferred to the epidermis during
the formation from the new integuments during the premolt stages.
The concentration in the carapace of shrimp from the earth pond
(batch P) showed contents of pigments comparable with those of the
three other treatment groups. The hepatopancreas was pigmented
much more than the epidermis and is similar to that of group A.
Compared to this "wild" batch, the control animals (0% carotenoid)
were de-pigmented and those fed with pellet enriched with 4% of
Pigmentech preserved the initial content of pigment, while those where
the feed enriched with 8% have the highest carotenoid level. In treatment
groups, the pigments from the alfafa concentrate appear to be better
absorbed than the natural pigments available in the earthen pond
resulting in a better colouration of the tissues.
The various forms of astaxanthin of which the quantity increases
in the epidermis and the hepatopancreas of supplemented shrimp
came from yellow carotenes and xanthophylls provided by the
supplementation in the feed.
Economical aspects
The alfalfa concentrate is also one of the least expensive sources of
carotenoids available on the market. If we compare the cost/g of active
ingredient, Pigmentech can be the most price competitive product
compared to most common products used as colour enhancers, such
as synthetic astaxanthin, paprika or the Spirulina based pigments.
In the above trials, we were only considering the effects on pigmentation
through the inclusion of carotenoids in the pelleted feeds. There are,
however, other additional benefits of the inclusion of the alfalfa-based
pigment as a source of limiting amino acids, essential fatty acids and
antioxidants, all of which have effects on economically important variables
such as growth and survival. Moreover the high level of protein in the
concentrate allow its use as an ingredient to substitute for other, more
expensive dietary components such as fish meal.
Conclusion
These studies showed that the incorporation of the natural alfalfa
concentrate at between 4 to 8% and fed during the last month of the
grow-out significantly increases the reddish colouration of shrimp after
cooking. Subsequent analysis showed a significant increase in
carotenoids, particularly in astaxanthin, in the shrimp tissue.
AQUA Culture AsiaPacific Magazine May/June 2006 | 35
FEED TECHNOLOGY
Table 2. Concentration in total carotenoids ((g) in the tissues of
the shrimp sample
