CULTIVATION OF SPIROGYRA FOR PAPER MAKING
PROJECT RFERENCE NO.:39S_BE_2269
COLLEGE
BRANCH
GUIDE
STUDENTS
: M S RAMAIAH INSTITUTE OF TECHNOLOGY, BENGALURU
: DEPARTMENT OF BIOTECHNOLOGY
: MR. M GOKULAKRISHNAN
: MS. SHWETHA UMESH
KEYWORDS:
Spirogyra, carbon, Nitrogen
INTRODUCTION:
Paper is an important commodity and will see an exponential rise in the years to come.
The sources of fibre are agro residues and forest which might not be enough for future needs of
paper production. The forest and agro residue need to undergo a lot of pre-processing for lignin
removal before the extraction is achieved. The widely used chemical process has been shown to
have just about 30-50% effectiveness of fibre extraction. With this brief background we see the
need of alternative sources for fibre extraction and production. Various different alternative
sources such have been studied widely, one such source is algae. With many distinctive
properties algae seems as a good fit for being the alternative source. Previously, people have
worked on using red algae, suspended micro-algae for fibre extraction and have observed good
yield and productivity. In my current study, I have selected Spirogyra as an alternative source
over other algae as it has suitable morphology and fiber properties for paper. Spirogyra is
filamentous in nature and has cellulose content similar to wood (17%) The main problem in using
algae as a source is its cultivation and harvesting. Hence in this study, I have focused on
Culturing spirogyra and harvesting them in a way suitable for paper industry.
OBJECTIVES:
1. Design a set up for culturing of spirogyra: A suitable set-up/ reactor design has to be
designed in order to facilitate continuous growth of spirogyra and also to make harvesting
of biomass easy.
2. Culture Spirogyra in different media to assess productivity: A suitable media has to be
selected for spirogyra growth, that supports high growth and at the same time has
minimum impact on nature.
3. Analyze various parameters to assess the performance of different set ups: Performance of
different set in terms of Nutrient cycling and complete utilization has to be estimated by
analyzing various basic parameters like pH, Carbon and nitrogen.
4. Assess harvestability of algae in a way suitable for paper production: Compare the
harvestability of different media and also the efficiency of design.
METHODOLOGY:
Algal samples from various different lakes and fresh water streams around Bangalore
were collected and tested. The collected samples were subjected to microscopic identification for
characterization of species distribution and selection of source with high number of Spirogyra
species.
The samples collected from site was washed thoroughly to remove any visible
contaminants like insects, waste attached to the fibrous algal body. Then later they were
transferred to a transparent plastic tub of dimension 30*36*15 cm filled with 10 L of water
supplemented with Urea and KH2PO4 to support their growth. The water was changed regularly
to avoid contamination from other algal species that grow in suspension and fibrous spirogyra
was washed each time with distilled water for a week. Once the growth of mother culture
stabilized it was allowed to further grow for a week to attain enough mass for sub culturing into
the different set- ups.
Algae cultures were cultivated in clear plastic tubs of dimension 30cm×36cm×15cm.
these tubs are transparent to allow passage of light. These were placed outdoor under natural
sunlight, temperature and environmental conditions. Algae was grown on mesh which were
placed 2cm below the water level as spirogyra grows in a fibrous manner on a surface near water
surface. The mesh that it used for growing spirogyra is of print screen quality steel mesh.
Spirogyra was cultivated in two different media set-ups in to compare growth and productivity:
Bold’s Basal Media and Soil set up. The cultures were maintained as duplicates.
Bold’s Basal Media:
Algae was cultivated in 7l of this media for a period of 10 days without any additional
nutrient in this growth period. Water Samples were collected from the tap at the bottom of the set
up so as to no disturb the culture. Water was lost on an everyday basis due to evaporation, hence
before sampling, the set ups were made up to 7 liters.
Soil set up:
Soil set up was chosen to imitate the open pond/ Agricultural field conditions. For soil set
up, 3 kg of clay soil was used in each reactor/set up, collected from paddy fields. The soil was
sun-dried before use. To this dry soil equal amount of water was added and allowed to settle for a
period of 3 hours following which the clumps in soil were broken manually, mixed and evened
out to a uniform layer and 7 liters of water was added.This culture was allowed to settle overnight
to allow complete sedimentation of soil particles. Similar to the Media tubs a mesh was placed 2
cm below the water level for algal cultivation and a tap was placed 1 cm above the soil layer for
water sample collection.After 24 hrs 1.2g of urea and 0.5 g of KH2PO4 was added to each soil
tub as a source for nitrogen and phosphorous.
After the overnight settling period of soil, nutrients were added to soil culture and both
the cultures were prepared for inoculation. 3g (dry weight) spirogyra was inoculated to each
culture and distributed on the mesh evenly and allowed to grow for a period of 10 days.In this
period the growth was monitored by collecting 100 ml water samples from each set up and
subjected to various analysis. The various parameters estimated during this phase were pH,
Carbonate- Bicarbonate by titrimetric method, free ammonia by distillation method followed by
titrimetric method and Nitrate by Sulphanilic acid method.
The mesh that algae grew on was rotated 3 cm everyday away from the water/ culture and
hence biomass was allowed to sun dry on the mesh. Over a period of 10 days a sheet of biomass
is expected to form on the mesh, such that it can be easily separated out from mesh and directly
sent for paper making by simply rolling up the sheets. Harvesting time and thickness was
estimated during the course of experiment.
RESULTS AND CONCLUSION:
Microscopic studies of cultures have shown that species contamination is present in both
the cultures but the species distribution varies widely. Synthetic media tubs had suspended microalgal species contamination, whereas soil tubs were contaminated with cyanophyceans.
Contamination with species is a common phenomenon when culturing in outdoor natural
conditions. It would pose a threat to the cultures only when the population of contaminant species
exceed that of desired species. In case of spirogyra cultivation the population of contaminant
species were low till 7th day after which they started increasing.
The pH of a system is important to monitor carbon and nitrogen. Apart from that pH is
also important for growth of biomass. Ideal pH for algal growth is 7 to 9.
Form the experiment it is observed that algae take up carbon mainly in the form of
carbonate and nitrogen in the form of free ammonia. The productivity and rate of uptake of
carbon and nitrogen are higher in soil set ups than in media set ups. Overall growth rate of
spirogyra is higher in soil tubs, supporting that soil cultures are more suitable and feasible for
algae culturing. Apart from growth rates and nutrient uptake, harvestability from different set ups
have showed that it is better in soil set ups than in media tubs and also that thickness plays a
major role in harvesting of biomass as a sheet without tearing.
Hence to conclude, we can say that soil set up supported better growth of algae and also
prevented nutrient losses compared to that of media set up. In terms of harvestability, uniform
sheet of algal harvest depends on the inoculum size and minimum thickness for a uniform harvest
is 0.2 mm.
FUTURE SCOPE:
Although the initial studies show a good growth and harvestability, large scale studies
would be necessary to understand the growth and feasibility for the technology for use. Work to
understand growth in different media will help us identifying different sources and potential use
of any waste water. A thorough study on the growth of algae so as to effectively grow algae to
obtain growth at the same time not compromise on the harvestability will be of utmost
importance especially with contamination lowering the yield. Spirogyra having good cellulose
quantity the property of cellulose plays an important role. Presence of crystalline cellulose makes
fiber properties useful for many purposes, a thorough study of the cellulose would be of
importance to make use of the fiber. A study on ability to pulp the spirogyra to obtain fibers with
minimal processing techniques would make the process all the more feasible and attractive to use.