Large Scale Production of Biofertilizers
Large Population of viable cells of effective strains of specific nitrogen fixing bacteria can be supplied through carrier based powder form of biofetlizer for cultivator use. Biofertilizers production technology includes isolation of bacteria, selection of suitable effective strain, preparation of mother or seed culture, inoculants isolation of bacteria, selection of suitable effective strain, preparation of mother or seed culture, inoculant production, carrier preparation and their mixing, followed by curing, packaging, storage and despatch.
The production of microbial inoculants of Rhizobium, Azotobacter and Azospirillum involves following steps except the both or liquid medium used is different for different organisms. The medium used for respective organism is as follows:
i) Rhizobium :Yeast Extract Mannitol
ii) Azotobacter : Ashby’s medium
iii) Azospirillium: Medium formulated by Okon et al. ( 1977)
iv) Phosphate solubilising bacteria: Pikiyskaya’s medium.
i) Preparation of Mother or Starter Cultures:
Starter cultures of selected strains are obtained after ascertaining their performance in green house and at field levels. The pure culture of efficient strain of nitrogen fixing organism is grown on respective agar medium on slant and maintained in the laboratory. A loopful of inoculum from the slant is transferred in a 250 ml capacity conical flask containing liquid medium. keep the conical flak on rotary shaker for 3-7 days depending whether they are fast growing or slow growing. The content of these flasks usually attain a load of 10 5- 10 6 cells per ml called mother culture or starter culture. This mother cultures are further multiplied in larger flasks.
2. Preparation of Broth Cultures:
Prepare liquid medium for respective organisms. Distribute equal quantity in big conical flasks (1000 ml). Sterilize it in autoclave for half an hour at 15 lbs pressure. After sterilization each flask containing suitable broth is inoculated with the mother culture in 1:5 proportions aseptically. Keep the flaks on rotary shaker for 96-120 hours until the viable count per ml reaches to 10 9 cells. The broths become more thick in consistency. This broth culture with population of 10 9 cells or ml should not be stored more than 24 hours or stored at 4 0C temperature.
3. Preparation of Carrier:
The carrier should have following characters:
a) It should have high organic matter above 60%.
b) Low soluble salts less than 1%.
c) High moisture holding capacity 150 to 200% by weight.
d) Provide a nutritive medium for growth of bacteria and prolong their survival in culture as well as on inoculated seed.
Lignite or peat is used as carrier in the preparation of Biofertilizers. The carriers are crushed and powdered to 200 to 300 mesh. Peat or Lignite powder is neutralized by addition of 1% calcium carbonate ( CaCO3) and sterilized at 15 lbs pressure for 3-4 hours in autoclave.
4. Preparation of Inoculate i.e. mixing:
The sterilized and neutralized lignite or peat is mixed with high count broth culture in galvanised trays. About 1 part by weight of broth is required to 2 part of dry carrier. Final moisture content varies from 40 to 50% depending upon quality of carriers.
5. Curing or Maturation:
After mixing the broth cultures and lignite or peat powder in 1:2 proportion in the galvanised trays then it is kept for curing at room temp ( 28 0C) for 5 to 10 days. After curing it is sieved to disperse the concentrated pockets of growth and to break the lumps.
6. Filling and Packing:
After curing, sieved powder is filled in polythene bagas of 0.5 mm thickness leaving 2/3 space open for aeration of the bacteria. Bag is weighted for desired quantity. Then the bag is packed by sealing. The polythene bag used for filling microbial inoculant should be printed with following information.
a) Name of Inoculants
b) Direction for use
c) Name of crops
d) Date of Manufacture.
e) Date of expiry.
7. Quality Checking:
Check viable count in the carrier based inoculants by dilution plate method at the time of manufactures. The viable cells count in the carrier based inoculants should be maintained as per ISI specifications.
The inoculants shall be stored by the manufacture in a cool place away from direct heat preferably at a temp of 15 0C and not exceeding 30 0C +- 2 0C for six months. For long survival of microorganisms the bag are stored in cold storage at 4 0C temp.