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  Section: General Biotechnology / Microbial Biotechnology
 
 
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Single Cell Protein (SCP) and Mycoprotein

 
     
 

Production of Algal Biomass

Algae (cyanobacteria and unicellular eukaryotes) grow autotrophically and synthesize their food by taking energy from sunlight or artificial light, carbon source from carbon dioxide, and nutrients from carbohydrates present in growth medium. In a few countries, cultivation of algae is carried -out in large trenches i.e. particularly in sewage oxidation ponds by using sunlight (Oswald, 1969) or in an artificial illumination conditions for use in life supportive systems for extended space exploration (Litchfield, 1967).

 

Chlorella strains are being used for a variety of applications in biotechnology. Due to their very high protein contents, they serve to improve protein deficiency and can be used as feed for production of animal protein. In many countries strains of Chlorella are utilized for sewage oxidation and waste water treatment (Kessler, 1989).

 

For cultivation of algae on sewage wastes, oxidation ponds are prepared, where sewage is allowed to accumulate. It is awaited till mixed cultures of algae grow (or inoculated with a singly prepared algal culture). For example, in Japan mixed culture of Chlorella ellipsoides and Scenedesmus obliguus was developed in open pond systems.

 

Factors Affecting Biomass Production

Following are the factors affecting the yield of biomass :

(i)    Illumination time;

(ii)   Light intensity;

(iii) Supply of CO2. Concentrations of CO2 differ in different conditions, for example, an alkaline lake. Lake Texcoco in Mexico, has high concentration of sodium carbonate. On the other hand, algal growth is limited as a result of liberation of CO2 and ammonia by bacterial activity;

(iv) Nitrogen sources (ammonium salts or nitrates are the suitable nitrogen sources which increase biomass yield);

(v)   Agitation of growing cells to maintain cells in suspension.

 

Biomass yield ranges between 12 and 15 g/m2/day (on dry weight basis) as obtained with Spirulina maxima and Scenedesmus quadricauda grown in out door pond conditions (Clement, 1968).

 

Mass cultivation of algae has been started in many countries, such as Japan, West Germany (now Germany), Mexico, Chechoslovakia, India, etc. In India, National Environmental Engineering Research Institute (NEERI), Nagpur has developed a technique of cultivating algae in sewage oxidation pond systems. This practice is also in use at NBRI (Lucknow), Hyderabad and other centres. Interestingly, experiments conducted at the CFTRI, Mysore have shown that the microalgae e.g. Scenedesmus acutus and Spirulina platensis could be cultivated on a large scale and used as food and feed as they are rich in protein and their nutrient value is comparable to conventional foods. A flow diagram of use of different groups of algae at various stages in waste water ponds and possible application of algal biomass is shown in Fig. 18.1.
 

Content

Advantages of producing microbial protein

Microorganisms use as single cell protein (SCP)

Substrates used for the production of SCP

Nutritional values of SCP

Genetic improvements of microbial cells

Production of algal biomass

 

Factors affecting bio­mass production

 

Harvesting the algal biomass

 

Spirulina as SCP, cultivation and uses

Production of bacterial and actinomycetous biomass

 

Method of production

 

Factors affecting biomass production

 

Product recovery

Production of yeast biomass

 

Factors affecting growth of yeast

 

Recovery of yeast biomass

Production of fungal biomass (Other than Mushrooms)

 

Growth conditions

 

Organic wastes as substrates

 

Traditional fungal foods

 

 

Shoyu

 

 

Miso

 

 

Sake

 

 

Tempeh  

Mushroom culture

 

Historical background

 

Present status of mushroom culture in India

 

Nutritional values

 

Cultivation methods

 

 

Obtaining pure culture 

 

 

Preparation of spawns

 

 

Formulation and preparation of composts

 

 

Spawning, spawn running and cropping

 

Control of pathogens and pests

 

Cultivation of paddy straw mushroom

 

Cultivation of white button mushroom

 

Cultivation of Dhingri (Pleurotus sajor-caju)

 

Recipes of mushroom

  Flow diagram of cultivation of algae in sewage oxidation ponds and possible application of algal biomass (modified after Vijayan, 1988).  
 

Fig. 18.1. Flow diagram of cultivation of algae in sewage oxidation ponds and possible application of algal biomass (modified after Vijayan, 1988).

 


Harvesting the Algal Biomass

Harvesting of algal culture becomes problematic because of settling down of cells at bottom and mixing of the algal cultures. The cells are recovered by concentration, dewatering and drying. Sometimes flocculants e.g. aluminium sulfate and calcium hydroxide and cationic polymers are added to the medium but they cannot be separated from the harvested cells. Therefore, this method warrants the application of SCP products in food and feed. Methods of separation and concentration also follow centrifugation, flocculation and centrifugation plus flocculation, but it is not economically feasible so far.

 

Harvesting the cyanobacteria, for example, Spirulina sp. is less troublesome as their spiral filaments float on the surface of water because of gas filled vacuoles in their cells which result in floating algal mats. Cells are able to fix atmospheric nitrogen. Algal mats are filtered and suspension of Spirulina is dried with hot air to get fine powder.

 

Algal yield from stabilization pond is around 114 tonnes/ha/year. In California 70 tonnes/ha/ year of Scenedesmus was obtained from sewage. From Bankok much high yield of about 170 tonnes/ha/year has been reported (Vijayan, 1988).

 

SCP Product of Spirulina

In 1821, Bernal Diaz del Castillo, for described the biscuits with the name 'tecuitlatl'. It contained dried mats of S. maxima collected from the Lake Texcoco. The biscuit was sold in the markets of Mexico. In 1964, J. Leonard (a Belgian botanist) took part in an expedition and noticed dried biscuits of blue green color in several village markets. The biscuit dike consumed by local population consisted of Spirulina platensis.

 

Many pilot plants for the production of Spirulina powder have been established in Japan, United States and European countries. Sosa Texcoco is the first Mexican Company to set up the first pilot plant in 1973 which produced about 150 tonnes Spirulina powder per annum; but the yield was increased to 1,000 tonnes in 1982. This company exported powder to the United States and prepared lozenges and capsules from the powders by adding vitamin A and C. The Mexican Company also supplied Spirulina powder to government institutions which were in charge of improving the nutritional situation of the population. Institutions used Spirulina powder to make biscuits and confectionery with a high protein content (Sasson, 1984).

Benefits from Spirulina SCP

Mass cultivation of Spirulatina offers several advantages over Chlorella and Scenedemus as given below:

(i)    Being a filamentous alga, Spirulina can be harvested by simple and less expensive methods such as nylon or cotton cloth filter.

(ii)   Filaments of Spirulina float on water surface due to presence of gas vacuoles. Hence, there is no problem of harvesting unlike Chlorella and Scenedesmus.

(iii) There is least chance of contamination in growth tanks of Spirulina as it grows at high alkaline pH 8-11.

(iv) Heat drying is sufficient for Spirulina as it has thin cell wall, whereas spray drying is required for Chlorella and Scenedesmus which is expensive,

(v)   Researches done by UNIDO programme in Mexico (1980), Mexican National Institute of Nutrition, Hyderabad (1988, 1990) under Indian Council of Medical Research on several aspects of possible adverse changes in multigeneration feeding tests on laboratory animals and humans have shown no adverse effects.

(vi) Spirulina is highly digestive (85-95%) due to thin wall and low nucleic acid contents (4%). It contains high percentage of digestible proteins (62-72%), vitamins, amino acids and other nutrients (Table 18.2). The aminogram of Spirulina is comparable to the FAD, milk and egg protein pattern.

 

Table 18.2. Composition of multin (i.e. dried powder of Spirulina fusiformis) (constituents are in per 100 of powder)*.

A. Major constituents (%)

 

C. Minerals (mg)

 

Total protein

64.6

Calcium

6.58

Fat

6.7

Phosphorus

977

Crude fiber      ;

9.3

Iron

44.7

Carbohydrates

16.1

Sodium

796

Calories

346

Potassium

1.28

B. Vitamins

 

D. Essential amino acids (%)

 

Beta - carotene

320,000IU

Lysin

2.99

Biotin

0.22 mg

Cystine

0.474

Cyanocobalamin (B12)

65.7 mg

Methionine

1.38

Folic acid

17.6 mg

Phenylalanine

2.87

Riboflavin

1.78 mg

Threonine

3.04

Thiamin

0.118 mg

 

 

Tocopherol

0.773 IU

 

 

* Analyzed by Michelson Laboratories Inc., California, USA (1988)

 

Mass Cultivation of Spirulina SCP

At present two types of farms for mass production of Spirulina SCP are under operation. A third type (i.e. enclosed system using transparent tube, biocoil or photobioreacter) is under development (Henrikson, 1990).

 

(i) Semi-natural lake system. Sosa Texcoco Lake (Mexico) and Lake Chand (Africa) offer an ideal environment for the natural growth of Spirulina. The product is expensive but of low quality due to contamination and pollution by uncontrolled natural factors. SCP of these lakes are good for fish and animal feed. Researches are in progress to refine the powder and make the products of good quality.

 

(ii) Artificially built cultivation system. The climatic conditions of most of the developing countries are such that favor mass outdoor production of Spirulina. Therefore, on the basis of water quality and nutrient status, this system can be grouped into the following two :

 

(a) Clean water system. This system is more expensive because of construction of artificial cultivation farms. These have shallow raceway ponds circulated with paddle wheels and high quality of nutrients. For the fast growth of Spirulina in clear water, addition of NaNO3 and NaHCO3 is necessary. pH of the water must be initially maintained to 8.5. It is a self pH adjusting alga which elevates the pH between 10-10.5. At this pH levels there is the least chance of contamination.

 

The Earthrise Spirulina Farm of California is the world's largest food grade Spirulina farm having the size of about 10 hectares with a capacity of production to about 120 tonnes/annum. The other big farms are operating in Japan, Thailand, Mexico, Taiwan, Israel, Vietnam, China and India.

 

In India, food grade Spirulina is cultivated at two main centres, one at Shri A.M.M. Murugappa Chettiar Research Central (MCRC), Madras, and the other at Central Food Technology and Research Institute (CFTRI), Mysore. Madras centre is the biggest food grade Spiulina farm in India. Its annual production capacity is of about 75 tonnes. The products are marketed in India and abroad as health food, baby food and multivitamin tablets.

(b) Waste water system. This system is applicable in highly populated countries like India where wastes are generated in high quantities and pose environmental problem. In this system, human and animal wastes and sewage are used for growth of Spirulina. The wastes are added into the digester to settle down the solid particles. The liquid effluent is used as a source of the nutrients and added in artificially constructed ponds. As desired NaNO3 and NaHCO3 are also mixed. S. platensis is found to grow better in sewage amended with NaCO3 and nutrients in different proportion and also in diluted sewage (Vijayan, 1988).

 

When full growth of Spirulina is over, it is screened from the pond and added to aquaculture to feed fish or dried in a small solar drier for human food.

 

This system is most suitable for third world countries where wastes are the major sources of pollution. R.D. Fox of France has developed the 'Integrated Health and Energy System Project' for poor countries to grow Spirulina and fight against the problems of food, malnutrition and environmental pollution. Today, a large number of these projects are in operation in the villages of developing countries like India, West Africa and South America. In India, the first integrated village system was established by Indo-France Govt. at Centre of Science for villages (CVS), Wardha (Maharastra).

 

The CVS distributes Spirulina cookies and nodules to malnourished children of local villages. This has shown very encouraging results.

 

Requirements for Growth of Spirulina

(i)      Algal tanks. Generally, circular or rectangular cemented tanks are constructed. The circular tanks are more preferred over the rectangular one because of ease in handling. Size may be according to convenience and yield needed. Depth should be about 25cm. Open tanks are suitable for tropical and subtropical regions.

(ii)     Light. Low light intensity is required at the beginning to avoid photolysis. Spirulina exposed to high light intensity is lysed.

(iii)    Temperature. Temperature for optimum growth should be between 35-40°C.

(iv)    pH. Spirulina grows at high pH ranging from 8.5 to 10.5. Initially, culture should be maintained at pH 8.5 which automatically is elevated to 10.5.

(v)    Agitation. Agitation of culture is very necessary to get good quality and better yield. The culture is agitated by brush, paddle power, pipe pumps, wind power, rotators, etc.

(vi)    Harvesting. The filaments of Spirulina float on surface of water forming thick mat. Therefore, it can be harvested by fine mesh steel screens, nylon or cotton cloths, etc.

(vii)   Drying. As it has a thin wall, sun drying is the most suitable and economical. Various trials done at CFTRI, Mysore and MCRC, Madras with sun drying have given good results.

(viii)  Yield. An average yield of 8-12 g Spirulina powder/m2/day has been obtained in India and other countries. This is equivalent to 20 tonnes /ha/annum. In warmer climate, the yield can increased to about 20 g/m2/day.

(ix)    Avoiding contamination. Although there is the least chance for contamination, yet regular monitoring of algal culture is necessary. Because the microbial load is likely to affect the quality and safety of the product. At MCRC and CFTRI the cultures of Spirulina were found either within or very close to safety limits of Indian Standard Institute (ISI) for baby food, to about 5xl05 propagules per gram (Venkataraman and Becker, 1985). Dried Spirulina powder is packed in aluminium bags or sealed in bottles and sent to market.

 

Uses of Spirulina Single Cell Protein

(i) As protein supplemented food. Since Spirulina is a rich source of protein (60-72%), vitamins, amino acids, minerals, crude fibers, etc., it is used as supplemented food in diets of under-nourished poor children in developing countries (Jeegi Bai and Seshadri, 1986; Sachan, 1991). The United Nations, Mexican National Institute of Nutrition, French Petroleum Institute and National Institute of Nutrition, Hyderabad have formulated four algal recipes as a weaning substitute for infants. In India, the Village Health and Energy System Projects are operated at CVS (Wardha). At MCRC, the products are distributed to the local under-nourished children. It has been found that 1 g of Spirulina tablets contains as much nutrition as one kg assorted vegetable.

 

(ii) As health food. Spirulina is very popular as health food. Most of Sosa Texcoco products are exported to USA, Europe and France where it is sold in health and food stores. It is the part of the diet of the US Olympic team. Jaggers take Spirulina tablets for instant energy. Since it provides all the essential nutrients without excess calories and fats, it is taken by those who want to control obesity. The MCRC has for the first time launched the project as health and baby food, and multivitamin powder and tablet under trade name 'Multin' and 'Multinal'.

 

(iii) In the therapeutic and natural medicine. Spirulina possesses many medicinal properties. Therefore, it is used as social and preventive medicine also. It has been recommended by medicinal experts for reducing body weight, cholesterol and pre-menstrual stress and for better health. It lowers sugar level in blood of diabetics due to the presence of gamma-linolenic acid and prevents the accumulation of cholesterol in human body (Nayak et al, 1988). It is a good source of (3-carotene (a precursor of vitamin A) and, therefore, helps in monitoring healthy eyes and skin. (3-carotene is known as the best anticancer substance (Schwartz et al., 1986). In 1989, UN National Cancer Research Institute announced that substances from blue-green algae are active against AIDS and cancer virus. In Vietnam its tablets are used to increase lactation in nourishing mothers.

 

(iv) In cosmetics. Spirulina contains high quality of proteins and vitamin A and B. These play a key role in maintaining healthy hair. Many herbal cosmeticians are making efforts to develop a variety of beauty products. Phycocyanin pigment has helped in formulating biolipstics and herbal face cream in Japan. These products can replace the present coaltar-dye based cosmetics which are known as carcinogenic.
 
     
 
 
     



     
 
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