Arthrospira (PROSEA)

Plant Resources of South-East Asia Introduction

List of species

Arthrospira fusiformis
- 1, habit of a trichome;
2, different forms of helicity.
A. maxima - 3, habit of a trichome.
A. platensis - 4, habit of a trichome.

Arthrospira Stizenb. ex Gomont

Protologue: Monogr. Oscill.: 246 (1892) [1893].

Family: Phormidiaceae

Chromosome number: Prokaryotic, thus no chromosomes

Arthrospira fusiformis (Woron.) Komárek & J.W.G. Lund, Algol. Stud. 58:11 (1990), synonym:

• Spirulina fusiformis Woron. (1934),
• Drouetian synonym: Porphyrosiphon notarisii (Menegh.) Kütz. ex Gomont (1892).

Arthrospira maxima Setch. & N.L. Gardner, Univ. Calif. Publ. Bot. 6(14): 377-379 (1917),

• synonyms: Spirulina maxima (Setch. & N.L. Gardner) Geitler (1932) not S. maxima C. Bernard (1909),
• S. geitleri G. de Toni (1935),
• Oscillatoria pseudoplatensis Bourr. (1970),
• Drouetian synonym: Microcoleus lyngbyaceus (Kütz.) P. Crouan & H. Crouan ex Gomont (1892).

Arthrospira platensis Gomont, Monogr. Oscill.: 247-248 (1892) [1893], synonyms:

• Spirulina jenneri (Hass.) Stizenb. var. platensis (Nordst.) ex Gomont (1892) [1893],
• S. platensis (Gomont) Geitler (1925),
• Oscillatoria platensis (Gomont) Bourr. (1970),
• Drouetian synonym: Microcoleus lyngbyaceus (Kütz.) P. Crouan & H. Crouan ex Gomont (1892).

The species in Arthrospira are often considered to be included in the genus Spirulina Turpin ex Gomont (1892) [1893]. Spirulina is, however, clearly different and phylogenically distant from Arthrospira.

In many papers on cultivated Arthrospira (often as "Spirulina") no species names are used. In these cases the material is usually considered to belong to A. platensis, although in some cases A. maxima material has been used.

• In most countries generally known as "spirulina"
• Thailand: sarai kleothong.

Arthrospira is commonly found in many alkaline salt lakes in Africa and America. It is cultivated in many (mainly) tropical countries, including South-East Asia. A. fusiformis is the name to be used for an autochtonous species found in tropical Asia and Africa. A. maxima appears to be essentially confined to Central America, while A. platensis seems to be more widely distributed and is mainly found in Africa, but also in Asia and South America. The name Arthrospira (Spirulina) platensis is commonly used for almost all commercially cultivated "Spirulina".

Early records show that dried A. maxima ("tecuitlatl") was consumed by natives in Mexico. In Chad a cake ("dihé") made of A. platensis is eaten with tomatoes, Capsicum peppers and various spices. About 9-13 g of this alga is consumed per meal in Chad, and it is eaten in 10-60% of the meals.

Arthrospira of food-grade quality is currently marketed (usually as "Spirulina") as powder or tablets with or without added calcium and/or vitamin C in the health food market ("nutraceuticals"). It can be directly used as food, mainly as substitute for green vegetables and is used especially for children. The mucoprotein cell walls of these algae are easy to digest. The food products marketed include protein powders with 10% Arthrospira, green coloured noodles or even chocolate bars containing Arthrospira. In Vietnam, the algal powder is mixed with milk and served as a nutritional supplement to treat malnourished children suffering from protein-deficiency diseases.

Several therapeutic effects have been recorded using Arthrospira. A cholesterol-lowering effect has been mentioned and Arthrospira is a potential food for persons suffering from coronary illness and obesity. Using the alga as supplement has been observed to produce hypocholesterolemic and hypoglycaemic effects, increased lactation in nursing mothers, chemoprevention and wound-healing properties. Antiviral and anti-cancer effects of Arthrospira and its extracts have also been found. Phycocyanin may have medical application. However, most claims have not been backed up by detailed scientific and medical research. No acute, chronic or subchronic toxicity of Arthrospira products has ever been detected.

Over 50% of the total production of Arthrospira powder is used as feed supplement. Especially in Japan Arthrospira is frequently incorporated as constituent of fish feed. It improves the palatability of this feed, the quality of the fish, enhances the colour of carp, and has health-promoting effects. Moreover, the mortality rate of fingerlings or post-larval stages in fish, molluscs and crustaceans can be reduced by 30-50% by adding 0.5-1.0% of Arthrospira to the feed. This also enhances growth of these animals. A comparable addition to fish feed can give a growth improvement of 17-25%. In Thailand and Vietnam Arthrospira powder is also used to feed silkworms.

Phycocyanin can be utilized as a natural pigment in the food, drug and cosmetic industries to replace the currently used synthetic pigments that are suspected of being carcinogens. The commercial product "Lina blue-A" is used as natural food colourant for confectionery, chewing gum, -icecream, dairy products and soft drinks. It does not react to light, but reacts slightly to heat. Another phycocyanin product obtained from Arthrospira, when modified to make it non-soluble in water, can be used as colourant in cosmetics. It does not run when it comes into contact with water or sweat, and is not a skin irritant. Small quantities of phycocyanin are used as a biochemical fluorescent tracer in immunoassays, microscopy and cytometry.

The carotenoids in Arthospira powder can cause pigmentation when used as a dietary supplement for cultured fish and shellfish such as koi carps, red tilapia, ayu, striped jack, and several kinds of prawns.

Waste-water treatment with Arthrospira offers many of the same advantages as treatment with other microalgae. The efficiency of ammonia stripping and phosphate removal from effluent by Arthrospira cultures needs further investigation, as does the use of these cyanobacteria in palm oil effluent in Malaysia and in waste water from tapioca factories in Thailand. The latter use, however, has already been applied commercially where 30 t of animal-feed Arthrospira biomass is produced annually.

Immobilized microalgae, including Arthrospira, are already used to remove heavy metals in commercially available products.

Commercial production of Arthrospira (usually as "Spirulina") at present is carried out on a large scale in Mexico, Taiwan, China, Thailand, Burma (Myanmar), United States (California), Japan and India. In 1993 there were 22 "Spirulina" commercial plants in the world, covering about 700 ha, producing 1000 t. By 1996 there were no less than 80 Arthrospira producers in China alone, mainly producing for export; the local market there is still quite small.

In Thailand, the Siam Algae Co. Ltd. cultivates Arthrospira in an inorganic medium in a pond area of 44 000 m² and the products are marketed as health food, animal feed and phycocyanin products. Production reached 100 t in 1980, 125 t in 1996 and will soon reach 150 t. Neotech Food Co. Ltd., Thailand, produced 36 t Arthrospira powder in 1996 from an area of 50 000 m² (30% human consumption, 70% animal feed).

The Myanma Pharmaceutical Industries in Burma (Myanmar) is the largest production site in the world based on natural blooms. In lake Twin Taung near Butalin, located in a volcanic crater, Arthrospira blooms occur mainly in native ponds covering about 130 000 m², resulting in about 32 t sun-dried algal powder for the local market.

In Vietnam, an Arthrospira plant of 5000 m² pond area of the National Mineral Water of Vinh Hao produces about 8 t of powder for export.

Arthrospira contains per 100 g dry weight: water 3-8 g, protein 50-66 g, lipids 4-10(-16) g, carbohydrates 13-25 g, fibre 4-10 g and ash 6-13 g, while the amount of usable protein in poultry and seafoods is considerably higher than in Arthrospira powder, it falls in the same class as most other meat and dairy products. In samples not washed sufficiently with acid water to remove absorbed carbohydrates, the ash content may be as high as 25%. The energy value averages 1680 kJ per 100 g. The net protein utilization values range from 41-63%, while digestibility is 83-84%. The proteins contain many essential amino acids (in % dry weight) including isoleucine (3.5-4.1), leucine (5.4-5.8), lysine (2.9-4.0), methionine (1.4-2.2), phenylalanine (2.8-4.0), threonine (3.2-4.2), tryptophan (0.91-1.1) and valine (4.0-6.0), as well as a high number of non-essential amino acids. The biological value of these microalgal proteins, however, is limited by the low levels of sulphur-containing amino acids. The levels of these amino acids can be raised by increasing the amount of sulphate in the culture medium.

The lipid fraction contains mainly hydrocarbons and terpenic alcohols, together with a low percentage of sterols. The major sterols in A. maxima are often clianosterol and cholesterol. The lipids contain, however, relatively high amounts of the polysaturated fatty acid 18:3 (ω-6) gammalinolenic acid. It forms 1.0-1.2% of the Arthrospira dry weight and 8-32% of the total fatty acids in these algae.

The algal powder is rich in vitamins such as β carotene, biotin, cyanocobalamin, pantothenic acid, folic acid, inositol, nicotinic acid, pyridoxine, riboflavin, thiamine and tocopherol. Mineral content in mg per 100 g of the product is: calcium 100-1400, chromium 0.2-0.3, copper 0.8-1.2, iron 47-150, magnesium 140-400, manganese 0.8-5.0, phosphorus 670-900, potassium 1330-1540, selenium about 0.04, sodium 27-900 and zinc 0.2-0.3.

Pigment content in mg per 100 g dry weight is: carotenoids 290-690, chlorophyll 600-1500, and phycocyanin 14-20. Of the carotenoids (in mg per 100 Arthrospira powder), especially β-carotene 50-140 and zeaxanthin 70-170 are interesting compounds.

• Trichomes solitary, 200-500 μm long, with very thin sheath, free-floating or showing gliding movement, multicellular, cylindrical, helical, loosely and regularly coiled, often slightly constricted at distinct cross-walls, apices slightly tapering or not.
• Cells with or without gas-vacuoles; terminal cell rounded or pointed, with or without a calyptra.
• Heterocysts absent.

A. fusiformis.

• Trichomes 3.4-6(-9.5) μm in diameter, not or only very slightly constricted, not or only slightly attenuate; helix very variable, up to 80 μm long, 15-50 μm wide, with end curves diminishing or widening intensely towards the ends.
• End cell rounded or slightly narrowed, occasionally with calyptra; all cells with regularly disposed gas-vacuoles.

A. maxima.

• Trichomes (6-)7-9(-10) μm in diameter, not or only very slightly constricted at the cross-walls, only slightly attenuate; helix rather long, 70-80 μm long, 40-60 μm wide, with end curves shorter than middle ones.
• End cell rounded, or with flat calyptra; all cells with regularly disposed gas-vacuoles.

A. platensis.

• Trichomes (4-)6-7(-8) μm in diameter, slightly constricted, attenuate, with rather short helix, 30-57 μm long, 26-36(-50) μm wide, with end curves not shorter than middle ones.
• Last 6-7 cells narrower; end cell longer than broad, rounded, capitate, with thickened outer wall; cells with or without irregularly disposed gas-vacuoles.

The development of A. platensis follows the common pattern of many other microorganisms which undergo a simple cell division without any sexual or differentiation step. A mature trichome breaks into small pieces through the formation of specialized cells known as necridia or lysing cells. The small trichomes are further fragmented to produce gliding, short chains consisting of 2-4 cells known as hormogonia. Each hormogonium can give rise to a new trichome, increasing in length and assuming the typical helical shape as the number of cells increases through binary fission.

Apart from autotrophic growth, A. platensis can also, in axenic cultures, be grown under mixotrophic and heterotrophic conditions. Some studies suggest that in mixotrophic conditions the autotrophic and heterotrophic forms of growth function independently in this alga, thus without mutual interaction. Heterotrophically grown cells (on glucose) have a lower pigment content than those in the autotrophic and mixotrophic cultures. Respiration-to-photosynthesis rates measured in axenic cultures are 1% at 20 °C and 4.6% at 45 °C. These rates are much lower than those recorded for outdoor cultures of A. platensis , where up to 34% of the biomass produced during the daylight period may be lost through respiration at night. However, the respiration rate is strongly influenced by light conditions during growth, and by light stress provoking photo-inhibition. A platensis cultures grown at less than the optimal temperature are more sensitive to photo-inhibition than those grown at the optimal temperature. In tropical countries, sub-optimal temperatures will occur during the early morning hours. However, lower temperatures result in decreased respiration, diminishing night loss of biomass in cultures. Nevertheless, a combination of relatively low temperatures and high light intensities may induce photo-inhibitory stress.

Exposure of cultures of A. platensis to high NaCl concentrations results in an immediate cessation of growth, followed by a usually somewhat slower growth rate. Photosynthesis and respiration activities both decrease markedly after exposure to high salinities. The salt-stressed cultures are very susceptible to photo-inhibition. Nevertheless, in China strains of A. platensis adapted to seawater are cultivated on a large scale in a seawater-based culture medium. The sand-filtered seawater is enriched with a commercial fertilizer (N:P:K = 12:12:12) and low concentrations of NaHCO₃ and Fe₂SO₄.

The taxonomy of blue-green algae in general is very complicated, and the same is true for Arthrospira. There is still much discussion about the separation and naming of species in this genus. A. fusiformis is often included in A. maxima. A new name was proposed, initially for the calyptrate specimens in A. fusiformis and later for the species name as a whole: A. indica T.V. Desikahary & W. Jeeji Bai. There is also much debate about the planktonic and non-planktonic nature of A. maxima and A. platensis respectively. The latter species, which forms massive water-blooms in tropical lakes, is considered to be non-planktonic by some specialists, while others think it well-founded to still consider A. platensis a planktonic species. The exact taxonomic position of the marine Arthrospira strains is also not yet clear.

Arthrospira grows abundantly in alkaline lakes characterized by high levels of carbonate and bicarbonate, and also in diverse habitats ranging from soils, marshes, brackish water, seawater, thermal springs and freshwaters. These algae are dominant organisms in alkaline lakes containing more than 30 g/l of salt (particularly sodium carbonate) and a pH close to 11. Optimal temperatures for growth range for different isolates from 30-42 °C, varying between 30-32 °C in selected temperate strains and 40-42 °C in tropical ones.

Zarrouk's medium is commonly used to culture Arthrospira. The medium contains high amounts of NaHCO₃ (16.8 g/l) as a carbon source and a buffer, and NaNO₃ (2.5 g/l) as a nitrogen source. High alkalinity is necessary for the growth of commercial Arthrospira , with a pH optimum of 8.3-11.0 for food growth. In outdoor pools a pH of 11.0 is growth-limiting. Usually Arthrospira is thought to be obligate photoautotroph and therefore unable to grow in the dark using organic carbon sources. However, it has been shown that heterotrophic growth of A. platensis is possible in axenic cultures. In laboratory conditions, the algae can be grown in shake flasks, aspirator bottles and photobioreactors.

Almost all commercial reactors for Arthrospira production are based on shallow raceways in which algal cultures sustained by a paddle wheel are mixed in a turbulent flow. Open raceways can be lined with concrete or formed as shallow earthen tunnels lined with PVC or some other durable plastic. In some cases, however, semi-natural lakes are used for Arthrospira cultivation. The size of commercial open raceway ponds for culturing Arthrospira ranges from 0.1-0.5 ha. Culture depth is usually maintained at 15-18 cm. In these ponds the optimal areal density of the algae must be maintained to prevent growth reduction due to self shading. This optimal areal density may also be affected by culture depth, the strain used, and the rate of mixing. Mixing (stirring) does not only enlarge the optimum density, but also prevents photo-inhibition caused by excess exposure to light of the algae staying too long in the upper and over-radiated water layers. It also lowers detrimental effects of too high oxygen concentrations in the upper layers of heavily photosynthesizing cultures. However, too high mixing and flow velocity result in fragmentation and increased coiling of the trichomes.

In an experimental plant in Thailand, the algae are mass-cultured in concrete raceway ponds (6 m × 26 m × 0.5 m) using waste water from a tapioca factory and stirred by paddle wheels. Another Thai experimental plant uses indigenous Arthrospira strains, mainly from north-eastern Thailand and capable of growing in brackish water. In India, in a rural location, Arthrospira is cultured using 2000 l tanks, agitated by manual stirring or by wheels driven by wind energy. In some cases a growth medium is used based on low-cost nutrients obtained from rural wastes such as bonemeal, urine, or the effluent of biomass digesters. In Vietnam, the medium is enriched with wastes from a fertilizer factory and the raceways are agitated by paddle wheels driven by wind energy. In order to use Arthrospira grown on wastewater as animal feed, it is necessary to minimize contamination of the biomass by choosing the right type of effluent to be treated. An integrated and sustainable approach is to combine the treatment of animal wastes with production of algal biomass for animal feed.

Arthrospira is also cultured in small-diameter transparent polyethylene tubes in Singapore, Italy and Israel. Other closed systems are flat plate reactors and biocoil facilities. Closed systems offer several advantages over open raceways: cultures are better protected from contaminants and can be effectively sterilized, while water loss and the ensuing increase in salinity of the medium are much reduced. They allow effective illumination due to a better surface-to-volume ratio and attainment of high biomass concentration. Because of much higher cell densities, areal volumes may be much smaller, thereby reducing harvesting costs. Finally, optimal temperatures may be established and maintained more readily in closed systems, resulting in higher output rates. All these developments still need to be tested on a large scale to evaluate whether the higher investment costs are indeed compensated by higher annual yields.

In China, most production plants are adopting a semi-closed culture system, where raceway culture ponds are covered by glass or transparent polythene sheet covering.

Contamination of Arthrospira by other microalgae can be prevented by maintaining a high bicarbonate concentration and by keeping the dissolved organic load in the culture medium as low as possible. Development of grazers in the cultures, mainly the amoeba type, can be arrested by addition of ammonia (2 mM). Ammonia (only 1 mM) can also be used to prevent further proliferation of Chlorella spp. in infested Arthrospira cultures. Contamination by green algae is often high during early cultivation stages, when the initial density of the Arthrospira inoculum is low. The amount of the contamination decreases as the Arthrospira builds up in density. Extra-cellular products of the Arthrospira may have some allopathic properties, which slow down the growth rate of Chlorella spp. Other contaminant algae are a blue-green Spirulina sp. and a green unicellular alga (Oocystis sp.). No cyanophages attacking Arthrospira have been observed so far.

Arthrospira can be easily removed from the medium by simple harvesting devices. The most used types of screens in filtration devices are inclining screens and vibrating screens. Inclining screens are 380-500 mesh with a filtration area of 2-4 m² per unit, capable of harvesting 10-18 m³ Arthrospira per hour. Biomass removal efficiency is up to 95% and two consecutive units are used for harvesting up to 20 l/m²/h, from which slurry (8-10% of dry weight) is produced. Vibrating screens can be arranged in double decks of screens up to 183 cm in diameter. These screens filter the same volume per unit time as the inclining ones, but require only one-third of the area and are very efficient.

In some plants in India, the biomass is harvested using twodeck gravity filters, consisting of two hemispherical cloth filters with filtration capacity of 200-670 l/m²/h. This technique is labour intensive and not very efficient, but well adapted to rural cultivation systems. In Madras (India), column gravity filters have been developed; the device consists of an eight-columned unit (1.55 m tall and 28 cm in diameter) with polyester cloth (30-40 μm pore size) and can continuously treat 5-10 m³ of culture/h. Yet another technique to remove Arthrospira biomass from the culture medium is vacuum filtration, a system good for small-scale production where a small pump with low energy consumption can be used.

In theory at least 40 g dry weight Arthrospira/m²/d can be obtained from a well-mixed outdoor pond, although 15-19 g/m²/d is the maximum reached so far in open systems, and usually less than half that figure is attained in commercial production plants. Daily productivity in polythenetube cultures ranges from 10-15 g dry weight/m²/d and productivity levels of 25-28 g dry weight/m²/d have been obtained in tubular bioreactors, while a similar areal productivity has been measured in flat plate reactors under natural illumination. In Thailand and China, however, the present productivity of Arthrospira in outdoor cultures is around 7-10 g dry weight/m²/d.

The algal slurry of Arthrospira (8-10% of dry weight) is rinsed in acid water (pH = 4) to remove the adsorbed carbonates. The biomass is stored at 0-20°C or frozen (18°C) before being sent for drying. Spraydrying is the most commonly used method of drying the biomass in commercial plants, although other systems, such as surface-drying (drums or plates), freeze-drying and sonic-drying are also used. Spraying yields a product which can be easily made into pills, while drumdrying yields a product in the form of flakes. The digestibility of spraydried products is usually higher than that of drumdried products. For optimal preservation and storage, moisture should not exceed 3-8%. The product, when stored under vacuum in sealed drums, can be kept for up to 4 years with little change in biochemical composition or nutritional properties.

Sun-drying is considered suitable if the product is to be used for animal feed. The biomass for feed is vacuumdried to 20% dry matter, mixed with different proportions of maize meal, and then dried in the sun. The resulting product consists of irregularly shaped pieces of Arthrospira/maizemeal mixture and is used as a fish feed.

In all cases a routine analysis of the chemical, physical, biochemical, and microbiological characters of the product is necessary in food-grade Arthrospira production, including bacteriological tests and checks for heavy metals and pesticides. Methods for chemical analysis procedures, preparation of growth media and suggested parameters for evaluation of the quality of Arthrospira powder are all well documented.

The consistency of biochemical composition of Arthrospira powder is remarkable, especially when the open nature of the production system is considered. Product quality and consistency will be even better in algae grown in closed bioreactors. The general appearance of Arthrospira powder should be uniform, without flakes and green to dark green, not brown. The taste ought to be mild; a bitter or salty taste may indicate insufficient washing or addition of preservatives. Arthrospira pills must have a smooth surface and no additional binders or coating should be used, because these may affect the digestibility of the product.

Many strains of Arthrospira are available from algal collections. Surprisingly few genetic studies have been done within Arthrospira. A mutant of A. platensis (Z19) is especially cultivated for the production of gamma-linolenic acid, while other mutants were induced for increased production of certain amino acids. A number of genes from Arthrospira have already been cloned and often have also been sequenced, which is a prerequisite for further genetic research.

Intensive research is being carried out to select highyielding strains of Arthrospira and to improve the culture systems so that productivity can be increased. It is expected that in the near future annual production will be doubled.

Commercial production of Arthrospira for health food is a lucrative industry. However, due to the strict requirements that have to be fulfilled in marketing foodgrade Arthrospira (as "Spirulina"), the focus has switched to the production of animal feed using agroindustrial wastes.

The main limitation of the present-day Arthrospira industry is the overall lack of sustainable productivity. With the increased demand for a high-quality product and a more sustainable and reliable production system for the future mass production of Arthrospira, development of closed systems is considered necessary. Much effort is required to further develop optimal closed photobioreactors.

Another approach is to develop technologies that are more suitable for local production of Arthrospira biomass in developing countries, and based on the use of conventional energy sources backed up by solar power. The operation of such a production process would be similar to that of an industrial plant, except that simpler methods would be used and more labour would be required. Plants need to be constructed using cheap local materials and inexpensive fertilizers and CO₂sources must be used. Appropriately designed high performance solar driers would help to cut production costs.

If considerable reduction in the costs of production units can be achieved, large-scale produced microalgal biomass has the potential to become a commodity traded in large quantities and not limited to the health-food market. It can then become an inexpensive high-protein supplement for human food and animal feed.

The use of closed culture systems and further selection of seawater-adapted Arthrospira strains will open up the possibility of using seawater with low bicarbonate concentrations, thus saving on the cost of water and medium. Moreover, the future development of integrated systems for Arthrospira biomass production and wastewater cleaning looks very promising.

The fluorescent properties of phycocyanin make it well suited for very diverse future commercial applications.

The Arthrospira-derived gamma linolenic acid cannot yet compete with seed oil as a dietary supplement. A silica gel method being developed to fractionate gamma linolenic acid from Arthrospira is showing promising results.

A polymer of potential use, poly-β-hydroxybutyric acid, occurs in A. platensis, where it accumulates during exponential growth to 6% of the total dry weight. The product can be used as a biodegradable thermoplastic polymer.

Selected strains of A. platensis can be used to bring about biotransformations to obtain valuable compounds. In India the alga has been used to biotransform codeine to morphine.

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Komárek, J. & Lund, J.W.G., 1990. What is "Spirulina platensis" in fact? Algological Studies 58. Fig. 2, p. 5. Redrawn and adapted by P. Verheij-Hayes.

• S.-M. Phang