Nutritional value of Chlorella vulgaris: Effects of
a Research Institute for the Biology of Farm Animals, Research Unit of Nutritional Physiology,Wilhelm Stahl Allee 2, 18196 Dummerstorf, Germany
b Institute for Cereal Processing Ltd. (IGV), Arthur-Scheunert-Allee 40-41,14558 Nuthetal-Rehbr¨ucke, Germany
Received 1 October 2004; received in revised form 14 February 2006; accepted 13 March 2006
Three processed products derived from the green algae C. vulgaris were investigated: (1) spray-
dried only (S-DA); (2) spray-dried and electroporated (ES-DA); (3) spray-dried; ultrasonicated treated(US-DA). A nitrogen-balance study was performed. Male growing Wistar rats, housed separately inmetabolism cages, were fed the three algal products as the sole protein source at 150 mg N per 100 gof body weight. A control group of rats was fed with casein at a level to give the same proteinnitrogen intake. The coefficients of total intestinal tract apparent crude protein digestibility for thedifferent C. vulgaris products were: S-DA = 0.47 ± 0.127% (mean ± S.D.), ES-DA = 0.44 ± 0.075%,US-DA = 0.57 ± 0.137%. Protein efficiency ratio was 1.4 ± 0.3, 1.0 ± 0.5 and 2.1 ± 0.3, respectively. N-balance was 41.86 ± 32.8 mg, 31.3 ± 17.3 mg and 66.7 ± 30.1 mg, respectively. The biologicalvalue was 93 ± 9.5%, 93.6 ± 10%, and 101 ± 5%, respectively. The coefficient of total intestinal tractapparent crude protein digestibility and biological value of C. vulgaris was enhanced by ultrasonictreatment and reduced by electroporating, thus ultrasonication may be a helpful technological processin practical processing of green algae in food industry. 2006 Elsevier B.V. All rights reserved. Keywords: Chlorella vulgaris; Nutritional value; Protein digestibility; Rats
∗ Corresponding author. Tel.: +49 38208 68666; fax: +49 38208 68652. E-mail address: (W.B. Souffrant).
0377-8401/$ – see front matter 2006 Elsevier B.V. All rights reserved. P. Janczyk et al. / Animal Feed Science and Technology 132 (2007) 163–1691. Introduction Chlorella vulgaris is a unicellular micro-alga that is ubiquitous in freshwater environ-
ments. The nutritional value of C. vulgaris was initially determined in 1950s–1960s Since then C. vulgaris has also been shown to have immune-modulating and anti-cancer properties Feeding micro-algae to elderly people or animals has been demonstrated to protect fromage-dependent diseases, particularly cardiac hypertension or hiperlipidemia (
The nutritive value of outdoor or indoor cultured C. vulgaris is of interest to the food
industry, especially in countries where the weather conditions do not allow massive cultureof higher plants. Nevertheless, first results of such studies were equivocal, depending uponthe technological process used to treat the algal mass. Different thermal processes appliedin order to destroy the robust cell wall, which restricts access of digestive enzymes to theintracellular components, also lead to destruction of amino acids and/or active substanceswithin the algal cells (In order to facilitate the access of digestive enzymes, and thus to enhance the nutritionalpotential of the cells, three different technological processes were applied in this study andtheir influence on the C. vulgaris protein digestibility and utilization was investigated in astudy undertaken in rats. 2. Materials and methods
The unicellular green algae C. vulgaris biomass was obtained from the Institute for Cereal
Processing Ltd. (IGV), Nuthetal-Rehbruecke, Germany, where it had been cultivated in aclosed photobioreactor PBR 4000 using sunlight (For feeding studies onanimals, three different types of algae have been used—spray dried (referred to subsequentlyas untreated), electroporated and ultrasonic treated.
The algal biomass was injected by a pump into the treatment cell with a delivery rate of
120 L/h (model HIS, self-made, TU-Berlin, Germany). The electrode gap in the cell was2 cm. A constant electrical field strength of 3 kV/cm was applied, this caused a specificenergy input of 7 kJ/kg algal biomass but no significant increase in temperature. Afterelectroporation the C. vulgaris biomass was spray dried. C. vulgaris biomass (start temperature of 20 ◦C) was treated using an ultrasonic device
(model UP 400S, Dr. Hielscher GmbH, Teltow, Germany). The pressure varied between
P. Janczyk et al. / Animal Feed Science and Technology 132 (2007) 163–169
Table 1Composition of spray-dried and technologically modified Chlorella vulgaris
2.0 and 5.6 bar, and the energy input between 961 and 1660 W. Even though the cell flowwas cooled with water as it passed through the ultrasonic device it became heated (to38.0–39.5 ◦C). It was therefore rapidly re-cooled to 20 ◦C before other treatments tookplace. The biomass was treated three times in total. Immediately after the third treatmentthe C. vulgaris biomass was spray dried. The composition of all green algae preparationsis shown in
2.4. Animals and experimental protocols
Male Wistar rats (Charles River Laboratories, Germany), were housed individually in
metabolism cages at 21.5 ± 1 ◦C, with a light regime 12 h light/12 h dark (Tecniplast, Italy). The metabolic cages allowed separate, and quantitative, collection of uneaten food, urineand feces. Rats were divided into four groups (n = 6) which received diets as summarizedin group fed with casein was an internal control group.
Two repetitions (an adaptation (7 days) followed by balance period (7 days)) were carried
out. The mean body weights were 135–136 g and 140–147 g at the beginning of the firstand second adaptation period, respectively. Each group of rats was fed the respective diet
a Protein source amount was calculated on nitrogen basis (150 mg N/10 g DM).
b N-free mixture consisted of 185 g cellulose, 371 g sugar, 148 g oil, 74 g vitamins (Vitamin A: 750 IE; Vitamin
B1: 1 mg; Vitamin B2: 1 mg; Vitamin B6: 0.5 mg; Vitamin B12: 0.5 mg; Vitamin C: 1 mg; Vitamin D3: 25 IE;Vitamin E: 2.5 mg; Vitamin K3: 0.1 mg; pantothenic acid: 1 mg; nicotine acid amide: 2.5 mg; choline hydrochloride:100 mg; folic acid: 0.1 mg; biotin: 0.01 mg; inositol: 12.5 mg; p-aminobenzoic acid: 5 mg; fulfilled ad. 1 g withwheat starch). A 148 g minerals (CaCO3: 68.6 g; Ca-citrate: 308.3 g; CaHPO4·H2O: 112.8; K2HPO4: 218.8;KCl: 124.7 g; NaCl: 77.1 g; MgSO4: 38.3 g; MgCO3: 35.2 g; Fe-ill ammoncitrate: 15.3 g; MnSO4·H2O: 0.2 g;CnSO4·H2O: 0.078 g; KJ: 0.041 g; NaF: 0.51 g; AlNH4(SO4)2·12H2O: 0.09 g; ZnCO3: 0.06 g) and 74 g wheatstarch/kg.
c Methionine was added to casein in the amount of 0.03 g casein. P. Janczyk et al. / Animal Feed Science and Technology 132 (2007) 163–169
in both adaptation and balance period according to their body weight (10 g per 100 g BW)and the diets contained 150 mg N in 10 g dry matter (DM). Feed intake was recorded andurine and feces were collected every day during the balance period. Hydrochloric acidwas added to urine and feces samples for storage until analyses were performed. Sampleswere weighed and the N-content and DM were determined. DM determination, in purealgae powder, feed, uneaten feed and fecal samples, was accomplished using the Weenderstandard procedure. N-content in all samples was analyzed in the elementary analyzer LECOCNS 2000 according to Dumas and was used for evaluation of N-balance. Amino acids weredetermined using automated amino acid ion chromatography (Biochrom 20 Plus Analyzer,Biochrom Ltd., Cambridge, UK). N-balance was used for calculation of coefficient of totaltract apparent crude protein digestibility (CTTAPD), amino acid digestibility (CTTAAD),net protein utilization (NPU) and protein efficiency ratio (PER). Endogenous and metabolicnitrogen were calculated using factors obtained in previous nitrogen-free experiments onrats, which had been done in the Institute. Using these data coefficient of total tract truecrude protein digestibility (CTTTPD) as well as biological value (BV) of crude protein wasthen calculated.
ANOVA followed by the Tukey HSD-test determined statistical significance of the data
(STATISTICA 6.0 software). A P-value <0.05 was considered significant. 3. Results
The results of the experiment concerning nutritional parameters are summarized in
Normal growth was observed in the internal control group (CAS), in which ratswere fed casein as protein source. Of the test treatments, the best growth was obtained inthe group fed ultrasonicated C. vulgaris (US-DA), the lowest growth rates were seen in thegroup fed electroporated algae (ES-DA). CTTAPD of the spray-dried algae was 0.47 ± 0.127(mean ± S.D.) which was similar to the electroporated C. vulgaris with 0.44 ± 0.075. CTTAPD of the ultrasonic treated algae 0.57 ± 0.137 was higher than both the other treat-ments but only significantly higher than the CTTAPD of electroporated micro-algae. ThePER was 1.4, 1.0 and 2.1%, respectively, and BV was 93.0 ± 9.8%, 93.6 ± 10.0% and100.7 ± 5.0%, respectively. CTTTPD was 0.53 ± 0.128, 0.51 ± 0.075 and 0.63 ± 0.138,respectively. 4. Discussion and conclusions
In our study rats fed ultrasonicated green micro-algae grew two times faster than rats fed
electroporated C. vulgaris and 1.5 times faster then rats fed untreated algae. The PER wasthe highest for the ultrasonicated C. vulgaris and was similar to PER of the micro-algaeobtained by hysical or chemical treatment of algal cells is necessary topermeabilise the cell wall, but not every process will increase the nutritional value of green
Table 3Nutritional parameters of differently processed Chlorella vulgaris (mean ±
* Different letters in columns show statistical difference by P<0.05. P. Janczyk et al. / Animal Feed Science and Technology 132 (2007) 163–169
algae, for example, high pressure homogenization decreased the digestibility of micro-algal protein (Similarly, in our experiments electroporation leads toless digestibility of algal protein. However, we also found that ultrasonication enhancesthe nutritional value of the green algae, which is clearly confirmed by all the measurednutritional parameters.
These results suggest that the technology of ultrasonication damages micro-algal walls
more efficiently than electroporation. The cell rupture is also more intensive than in thespray-dried green micro-algae. It may also be that electroporation not only damages the cellwall, but also intracellular structures and molecules, hence reducing the nutritional valueof the micro-algae. These observations have great practical value for the use of the greenmicro-algae as a food component and clearly show different effects of the three technologicalprocesses on C. vulgaris preparations with respect to it’s potential as a biological proteinsource. Acknowledgments
The investigations were performed in collaboration with “Frankenf¨order Forschungsge-
sellschaft mbH” and with financial support from the Ministry of Economics of the StateBrandenburg. References
Justo, G.Z., Silva, M.R., Queiroz, M.L.S., 2001. Effects of the green algae Chlorella vulgaris on the response of the
host hematopoietic system to intraperitoneal Ehrlich ascites tumor transplantation in mice. Immunopharmacol. Immunotoxicol. 23, 119–132.
Komaki, H., Yamashita, M., Niwa, Y., Tanaka, Y., Kamiya, N., Ando, Y., Furuse, M., 1998. The effect of processing
of Chlorella vulgaris: K-5 on in vitro and in vivo digestibility in rats. Anim. Feed Sci. Technol. 70, 363–366.
Konishi, F., Tanaka, K., Himeno, K., Taniguchi, K., Nomoto, K., 1985. Antitumor effect induced by a hot water
extract of Chlorella vulgaris (CE): resistance to Meth-A tumor growth mediated by CE-induced polymor-phonuclear leukocytes. Cancer Immunol. Immunother. 19, 73–78.
Konishi, F., Tanaka, H., Kumamoto, S., Hasegawa, T., Okuda, M., Yano, I., Yoshikai, Y., Nomoto, K., 1990.
Enhanced resistance against Escherichia coli infection by subcutaneous administration of the hot-water extractof Chlorella vulgaris in cyclophosphamide-treated mice. Cancer Immunol. Immunother. 32, 1–7.
Konishi, F., Mitsuyama, M., Okuda, M., Tanaka, K., Hasegawa, T., Nomoto, K., 1996. Protective effect of an
acidic glycoprotein obtained from culture of Chlorella vulgaris against myelosuppression by 5-fluorouracil. Cancer Immunol. Immunother. 42, 268–274.
Lin, Y.Ch., 1969. The supplementary effect of algae on the nutritive value of soybean milk. JFMA 68, 15–21. Lubitz, J.A., 1963. The protein quality, digestibility, and composition of micro-algae, Chlorella 71105. J. Food
Morimoto, T., Nagatsu, A., Murakami, N., Sakakibara, J., Tokuda, H., Nishino, H., Iwashima, A., 1995. Anti-
tumour-promoting glyceroglycolipids from the green algae Chlorella vulgaris. Phytochemistry 40, 1433–1437.
Noda, K., Ohno, N., Tanaka, K., Kamiya, N., Okuda, M., Yadomae, T., Nomoto, K., Shoyama, Y., 1996. A
water-soluble antitumor glycoprotein from Chlorella vulgaris. Planta Med. 62, 423–426.
Okamoto, K., Iizuka, Y., Murakami, T., Miyake, H., Suzuki, T., 1978. Effects of Chlorella alkali extract on blood
pressure in SHR. Jpn. Heart J. 19, 622–623.
Pulz, O., Scheibenbogen, K., Gross, W., 2000. Biotechnology with cyanobacteria and microalgae. In: Rehm, H.-J.,
Reed, G. (Eds.), Biotechnology, vol. 10, 2nd ed., Wiley-VCH, Weinheim, pp. 105–136. P. Janczyk et al. / Animal Feed Science and Technology 132 (2007) 163–169
Saleh, A.M., Hussein, L.A., Abdalla, F.E., El-Fouly, M.M., Shaheen, A.B., 1985. The nutritional quality of drum-
dried algae produced in open door mass culture. Z. Ernahrungswiss. 24, 256–263.
Sano, T., Tanaka, Y., 1987. Effect of dried, powdered Chlorella vulgaris on experimental atherosclerosis and
alimentary hypercholesterolemia in cholesterol-fed rabbits. Artery 14, 76–84.
Sano, T., Kumamoto, S., Kamiya, N., Okuda, M., Tanaka, Y., 1988. Effect of lipophilic extract of Chlorella vulgaris
on alimentary hyperlipidemia in cholesterol-fed rats. Artery 15, 217–224.
Singh, A., Singh, S.P., Bamezai, R., 1999. Inhibitory potential of Chlorella vulgaris (E-25) on mouse skin papil-
lomagenesis and xenobiotic detoxication system. Anticancer Res. 19, 1887–1892.
Tanaka, K., Konishi, F., Himeno, K., Taniguchi, K., Nomoto, K., 1984. Augmentation of antitumor resistance by
a strain of unicellular green algae Chlorella vulgaris. Cancer Immunol. Immunother. 17, 90–94.
Tanaka, K., Koga, T., Konishi, F., Nakamura, M., Mitsuyama, M., Himeno, K., Nomoto, K., 1986. Augmentation
of host defense by a unicellular green algae, Chlorella vulgaris, to Escherichia coli infection. Infect. Immun. 53, 267–271.
Tsuchida, T., Mashiko, K., Yamada, K., Hiratsuka, H., Shimada, T., Itagaki, Y., Fujinuma, H., Samejima, K., Naka-
mura, T., Hasegawa, T., Matsubayashi, T., 2003. Clinical study of gamma-aminobutyric acid-rich Chlorellafor subjects with high-normal blood pressure and mild hypertension. J. Jpn. Soc. Nutr. Food Sci. 56, 97–102.
Yasukawa, K., Akihisa, T., Kanno, H., Kaminaga, T., Izumida, M., Sakoh, T., Tamura, T., Takido, M., 1996.
Inhibitory effects of sterols isolated from Chlorella vulgaris on 12-0-tetradecanoylphorbol-13-acetate-inducedinflammation and tumor promotion in mouse skin. Biol. Pharm. Bull. 19, 573–576.
Therapeutic Use Exemption (TUE) PROCEDURE 2009 Table of contents 1. Introduction and Scope 2. Terms and Definitions 3. TUE Application Process 4. Asthma 5. Criteria for Granting a TUE 6. TUE Committee 7. Confidentiality of Information 8. Declaration of Use Process 9. Clearinghouse 10. Taking effect Art. 1: Introduction and Scope One of the principles of anti-doping policies i
New Patien Patien t Qu What is your main reason for seeing us today? Previous Diagnosis: Have you received a diagnosis from a previous pain specialist? Did he/ she explain the cause or reason for the pain you're feeling? Was your Injury on the job/ auto accident/ fall/ spontaneous occurance/ sports injury/ other:Is the pain constant/ intermittent/ sporadic/ worse at night/ worse