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Slide 1

High Protein and β-Carotene Yield of Dunaliella spp. Exposed to High Light Intensities and Nutrient Limitation

Samia K. Mikhail & Wagdy LabibNational Institute of Oceanography and Fisheries,Kayet Bey, Anfoshi, Alexandria, Egypt

Slide 2

  • This study was done to test some algae for the highest dry weight, protein, essential amino acids and carotene contents in order to use its dried form as a mixture with boiling soybean meal, to feed Oreochromus niloticus fry.

  • Soybean meal is commonly utilized as plant protein in fish feeds, because it is readily available, palatable, inexpensive and has balanced amino acid profile and high nutritional value. However, soybean meal when compared to fish meal is characterized by a lower composition of essential amino acids, mainly methionine, lysine and threonine.

Slide 3

  • Under non inducing non accumulating conditions D. salina cells are green and contain only 0.3% of β-carotene

  • Under extreme environmental conditions (high salinity, high temperature, high light and low nitrogen supply) concentrations up to 14% of dry weight have been reported and the cells appear orange-red.

Stress

Slide 4

  • Dunnalilla spp. is cultivated commercially in large outdoor ponds and harvested to produce high β- carotene dry algal meal and concentrated algal β- carotene in oil, used by the health food industry and food coloring.

  • Dunaliella pigments have great commercial value as natural colorants in cosmetics and pharmaceutical industry, besides their health benefits.

  • Dunaliella capsules are now commonly prescribed health foods for improving vitality and longevity of human beings.

Slide 5

  • The study samples were collected from the water near El-Mex

  • Saline Company west of Alexandria, Egypt, which contains salt

  • marches.

  • The commercial salt present with this algae, is considered as

  • impurity.

  • The aim of work is not only to remove these impurities but

  • also make use of it in industry.

Slide 6

Material and Methods

  • Cells were cultivated separately in both MH modified artificial medium and enriched natural medium (KNO3, KH2PO4 and Fe-EDETA) pH 8.0. Salinity was adjusted to 35 ‰.

  • Dunaliella spp. were cultured in both two media and incubated at 28ºC± 1ºC in an incubator with vigorous hand agitation once a day, to prevent clumping of the cells, under continuous illumination at 4 light intensities of 7.5x103, 12.5x103, 17.5x103 & 22.5x103 Lux and 7.5x103 Lux at 12L: 12D photoperiod.

  • The cells were washed twice with nitrate-free medium and incubated for 2 days.

  • Nitrate-starved cells were cultured in the two media with different nitrate concentrations, as KNO3, [0.1, 0.4, 1.0, 6.0 & 12.0 mM] under continuous light at 28± 1ºC.

Slide 7

  • Samples from early stationary–phase were harvested and the different biochemical parameters were measured.

    • The cell concentration was calculated by cell counting.

    • Chl. a and β-carotene were determined Eijckelhoff & Dekker (1997).

    • The total nitrogen content was determined by the micro-Kijeldahl method and the protein content was calculated by multiplying the value of total nitrogen by 6.25.

  • In order to overcome the lake of Dunaliella spp. at times other source as macophytes, marine and freshwater algae (some Chlorophyta, Rhodophyta, Cyanophyta and Euglena spp.) were tested for their protein content and EAAS using amino acid analyzer.

  • Slide 8

    Results and discussion

    D. bardawil

    D. salina

    • The data showed the highest value of cell density, chlorophyll a and β-carotene for the tested algae (D. salina and D. bardawil) cultured in both media was observed with the onset of the stationary phase at the 8th day.

    • The maximum cell density for D. salina (11.47 x 106 cells ml-1) and Chl. a content (54.64µg ml-1) were higher than that of D. bardawil (1.47 x 106 cells ml-1& 15.64 µg ml-1).

    Slide 9

    β-carotene content

    • For all results Natural media gave better values than the synthetic one for both species.

    • This was clear with β-carotene content for both species.

    • D.bardawil yields higher content of β-carotene (15.43 & 13.04 µg ml-1) than D. salina (14.77 & 12.31 µg ml-1) in both media and natural media was always higher.

    D. Bardawil (N.M, 15.43 & S.M, 13.04 µg ml-1)

    D. Salina (N.M, 14.77 & S.M, 12.31 µg ml-1)

    Slide 10

    Effect of different light intensities

    I- Total protein

    • The present study showed

    • A stimulating effect of light intensities on total protein of Dunaliella spp.

    • A gradual increase in protein value with increasing light intensity

    • Protein contents in natural media (maximum of 86.2 % at 22.5

    • x 103 Lux) were higher comparing with that in synthetic media

    • (82.4%).

    Slide 11

    • Diurnally grown cells of D. salina had lower protein content than that of cells growing similarly but under continuous light in synthetic and natural media.

    Slide 12

    II- Dry weight

    • Dry weight gained by the tested algae Dunaliella spp. Showed also gradual increase by increasing light intensity in both media.

    • Maximum elevation in dry weight was observed at the highest light intensity of 22.5x103.

    • N.M gave better results.

    Slide 13

    III- β-carotene content

    65.37

    Under stress conditions transient response in the surviving population of Dunaliella spp., including rounding of the cells, decreased motility, change in colour from green to deep orange and cessation of cell division was observed.

    58.43

    5.27

    4.51

    • The β-carotene content per cell increased with light intensity in both species.

    • The maximum content for D.salina was 5.27 (N.M) and 4.51 (S.M), pg cell-1.

    • D. bardawil have higher values 65.37 (N.M) and 58.43 (S.M) pg cell-1.

    Slide 14

    Effect of nitrogen depletion I- carotene content

    • The carotene content changed markedly with increased limitation by nitrate, in the nitrate starved cells in both media carotene content reached 56.52 (N.M) and 40.01 µg ml-1 (S.M) for D. salina and 348.58 (N.M) and 330.71 µg ml-1 (S.M) for D. bardawil.

    • Growth and chlorophyll synthesis slow down due to nutrient depletion and algae cease to divide.

    Slide 15

    II- Protein contents

    • A parallel relationship between the nitrogen concentration in the media and protein content of Dunaliella spp. was observed in the two media.

    • In contrast with carotene accumulation, protein content was higher in D. salina and natural media was better with protein production for both species.

    Slide 16

    • Parsons et al. (1961) suggested that a protein, carbohydrate and fat ratio of approximately 4:3:1 is suitable for zooplankton nutrition.

    • In our data this ratio is true for protein: fat ratio of D.salina cultured under high light intensity 22.5 x103 Lux in both natural and synthetic media, but not true for carbohydrate ratio.

    • Dunaliella spp. with high source of carbohydrate content will be a good diet for animal feeding.

    Slide 17

    Essential amino acids

    • The EAA, mainly methionine & valine are relatively higher in D. salina.

    • Histidins is highest in D. bardawil.

    • A mixture of

      Dunaliella spp. is a good source for the three EAA.

    • Other algal species were tested for using as supplementary food for mix/replacing D.salina as dried algae with BSB in the mixture diets.

    Slide 18

    Effect of replacement of fish meal protein with boiled full fat soybean seeds with dried algae on growth performance, nutrient utilization and some blood parameters of Nile tilapia (Oreochromis niloticus)Raky F. Attalla1, Samia K. Mikhail2National Institute of Oceanography and Fisheries1: Inland water, Cairo, Egypt.2: Kayet Bey, Anfoshi, Alexandria, Egypt.

    Slide 19

    • Mixture of boiled full fat soybean seed and dried algae (Dunaliella spp.) at mixing level of 1:1 was tested to replace fish meal protein on Nile tilapia fry at

      - 50% (soybean) + 50% FM

      - 50% mixture + 50% FM

      - 75% mixture + 25% FM

      - 100% mixture

      - 100% soybean

      Results obtained are summarized as follows:

    • 50 and 75% levels released no significant effects on final weight; average weight gain ; daily gain and survival rates compared to the control groups (100% FM) while the same parameters were decreased significantly in the other treatment groups.

    • Mixture at 50% level had insignificant effects on feed conversion ratio and protein efficiency ratio compared to the control groups while the same parameters were negatively affected in the other group.

    Slide 20

    Conclusion

    • Natural media gave better values than the synthetic one for both species.

    • Diurnally grown cells of Dunaliella spp. had lower values than cells growing similarly but under continuous light in synthetic and natural media.

    • The data showed the highest value of cell density, chlorophyll a and β-carotene for the tested algae (D. salina and D. bardawil) cultured in both media was observed with the onset of the stationary phase at the 8th day.

    • D. salina showed higher cell density and Chl. a content than that of D. bardawil.

    • D.bardawil yields higher content of β-carotene than D. salina

    • The data showed a gradual increase in protein, dry weight β-carotene content value with increasing light intensity

    Slide 21

    • The carotene content changed markedly with increased limitation by nitrate, higher in D. bardawil

    • In contrast with carotene accumulation, protein content showed a parallel relationship with nitrogen concentration and was higher in D. salina

    • The EAA, mainly methionine & valine are relatively higher in D. salina. Histidins is highest in D. bardawil. A mixture of Dunaliella spp. is a good source for the three EAA.

    • Dunaliella spp. with high source of carbohydrate content will be a good diet for animal feeding.

    • up to 50 % treated Soya and dried algae can be incorporated safely and economically in diets of Nile tilapia.

    Slide 22

    Thankyou

    Dr. Samia Kamel

    National Institute of Oceanography & Fisheries

    (NIOF)

    E-mail: Mikhail_Samia@Yahoo.com


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