Ultrahigh productivity photobioreactors for algal biofuel production

308 Pages·2014·17.48 MB·English

by  Kirsty Barber

Ultrahigh productivity photobioreactors
for algal biofuel production

Kirsty R. Mokebo

A thesis submitted for the degree of Doctor of Philosophy

Department of Chemistry
University of Bath
August 2012

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Acknowledgements

Firstly, I would like to thank my supervisor Professor Matthew Davidson for giving me the
opportunity to work on this project, for his support, guidance and enthusiasm throughout
my PhD. I would also like to thank my various co-supervisors over the course of my PhD,
Drs Alexei Lapkin, Pawel Plucinski and Laura Torrente Murciano particularly for their
guidance with the engineering aspects of this work.

The University of Bath are acknowledged for funding this PhD and edf and Knowledge
West for funding the building of the photobioreactor.

Many thanks to Paul Frith in the Faculty of Science Engineering workshop for his
technical knowledge, and Michael Thomas of the Electrical Engineering department for his
attention to detail, for the illuminated section of the photobioreactor. Thank you also to
Robert Brain, Fernando Acosta, John Bishop and Richard Bull of the Chemical
Engineering department for their help and support with my requests for practical assistance
and specific parts for the photobioreactor. Thanks to Phil the glass for his assistance too.

I would like to thank Dr Chris Chuck for his enthusiasm and encouragements, and to Dr
Konstantin Loponov for his preliminary designs of the photobioreactor. Dr Chris Bannister
is gratefully acknowledged for his assistance with the design of experiments work and flow
dynamics calculations. Thank you to the students who have worked on this project, Akid,
Andrew, Becca, Charlotte, Lu, Matthew and Phatra.

Many thanks to the Biology department, particularly Holly Smith-Baedorf, we got there in
the end! Thanks also to Professor Rod Scott, Philippe Mozzanegga and Dan Murray.
Thanks to Carlo Di Iulio and Dr Matthew Jones for the use of the GCMS, thanks to the
Davidson, Jones and Chuck group for the high lab spirit and friendship throughout my PhD!
My PhD wouldn’t have been the same without Anna, Becca, Ben, Carlo, Cathy, Chris,
Chris, Dan, Daniel, Emma, Holly, Justin, Lisa, Lois, Rhod and Tom – thank you!

Thank you to my mum and dad for their encouragements and support, to Joe, Michelle and
Shaun and to all at Living Hope Church.
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Abstract

Algal biodiesel is a biodegradable and sustainable alternative to traditional petroleum fuels.
Algal biodiesel is synthesised from algal lipids via transesterification and has many
desirable physical properties for fuel use. Current photobioreactors are inefficient. This
thesis looks to increase efficiency and reduce energetic running costs. This was undertaken
by the design, construction and trialling of an LED photobioreactor. The controlled growth
of the algae, specifically Chlorella emersonii, using pulsed monochromatic or bi-chromatic
light conditions with comparison to continuous white light to improve light economy is
explored in this thesis. The prediction of biodiesel profile from the growth conditions is
also investigated for Chlorella emersonii.

Chapter 1 is a general introduction to the area of algal biodiesel. This introductory chapter
reviews the current literature regarding microalgae growth conditions and control,
processing microalgae to produce biodiesel and photobioreactor designs for the controlled
growth of algae. The known effects of different light sources and types on algal growth are
also reviewed.

Chapter 2 concerns the pulsing-LED vertical airlift photobioreactor design, construction
and testing, including an overview of the system constructed and the process of design to
combat specific issues. Results from the testing of the photobioreactor are reported in this
chapter which include analysis of the resultant fatty acid methyl ester (FAME) profile of
algae grown under various pulsed mono-chromatic and bi-chromatic light conditions and
the comparison to continuous white light. This chapter draws together the hypotheses and
stand-alone observations reported in the current literature allowing direct comparisons for
different light conditions and conclusions to be reported which include the effect on
resultant FAME profile and not just lipid percentage.

Chapter 3 explores the effect of environmental factors on the fatty acid methyl ester
composition of the algal biodiesel. This chapter describes the effect of carbon dioxide,
nitrate, phosphate and iron levels, length of culture and the effect of supplementary carbon
sources on Chlorella emersonii growth and resultant FAME composition. The result of
synergetic effects of nutrient levels and length of algal cultivation are analysed in addition
to the stage of algal growth and its impact on FAME profile.
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Chapter 4 details the procedures used for the growth of algae, the production of the algal
biodiesel and the development of techniques used for analysis of the resultant biodiesel.
The techniques and conditions employed for the growth of the algae as well as the
extraction and transesterification of the algal lipids are explained.

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Contents Page

1 Introduction .................................................................................................................... 1
1.1 Preamble .................................................................................................................. 1
1.2 Biodiesel .................................................................................................................. 1
1.2.1 The requirement for biofuels as a transitional technology ............................... 2
1.2.2 Fatty acid methyl ester (FAME) profiles and fuel properties of biodiesel ...... 3
1.2.3 Issues with first generation biofuels ................................................................. 5
1.2.4 Potential of algal biofuels................................................................................. 7
1.2.5 Problems with algal biofuels .......................................................................... 10
1.2.6 Commercial ventures in algal biofuels ........................................................... 11
1.2.7 Policies and government goals ....................................................................... 11
1.2.8 Economical cost of algal biofuel .................................................................... 12
1.2.9 Energy costs and considerations .................................................................... 14
1.2.10 Transesterification .......................................................................................... 15
1.3 Microalgae ............................................................................................................. 17
1.3.1 Microalgal species .......................................................................................... 17
1.3.2 Nutrients ......................................................................................................... 19
1.3.3 Growth parameters ......................................................................................... 20
1.3.4 Growth rates ................................................................................................... 20
1.3.5 Algae growth conditions ................................................................................ 21
1.3.6 Effect of carbon sources on algal growth ....................................................... 22
1.3.7 Algal growth: the interrelated effects of carbon dioxideand pH ................... 22

1.3.8 Mixotrophic algae growth .............................................................................. 24
1.3.9 Salinity effects on algae growth ..................................................................... 25
1.3.10 Effect of limiting conditions on lipid content of algae .................................. 25
1.3.11 α-tocopherol and carotene .............................................................................. 29
1.4 Downstream processing and extraction of lipids from algae ................................ 30
1.4.1 Separation of algae from growth media ......................................................... 30
1.4.2 Lipid extraction utilising solvents .................................................................. 32
1.4.3 Alternative extraction techniques ................................................................... 33
1.4.4 Fatty acid methyl ester profile analysis .......................................................... 36
1.5 Photobioreactors .................................................................................................... 37
1.5.1 Considerations for photobioreactor design .................................................... 37
1.5.2 Algal reactor types ......................................................................................... 40
1.5.3 Examples of photobioreactors ........................................................................ 42
1.5.4 Mixing ............................................................................................................ 43
1.5.5 Carbon dioxide supply and control ................................................................ 45
1.5.6 Oxygen saturation .......................................................................................... 46
1.5.7 Mass flow control meter................................................................................. 47
1.5.8 Cleaning ......................................................................................................... 47
1.5.9 Light requirement and light emitting diodes .................................................. 47
1.5.10 Photosynthesis ................................................................................................ 49
1.5.11 Effect of light on algae growth and productivity ........................................... 52
1.5.12 Photoinhibition in photosynthesis .................................................................. 55
1.5.13 Light regimen effects ..................................................................................... 56
1.5.14 Effect of pulsed light on algal growth and acclimatisation ............................ 56
1.5.15 Effect of wavelength on algae growth ........................................................... 62
1.5.16 Carbon dioxide capacity and affinity in photosynthesis ................................ 63
1.6 Summary and aims ................................................................................................ 64
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1.7 References ............................................................................................................. 66

2 Photobioreactor design, construction and testing......................................................... 81
2.1 Initial design ideas ................................................................................................. 82
2.2 Overview of photobioreactor ................................................................................. 83
2.3 Construction materials ........................................................................................... 83
2.4 Airlift section ......................................................................................................... 86
2.4.1 Sparger ........................................................................................................... 91
2.5 Illuminated section ................................................................................................ 92
2.5.1 Temperature and pH logging ......................................................................... 97
2.5.2 Cooling ........................................................................................................... 97
2.5.3 Light emitting diodes illumination ................................................................. 98
2.6 Mass flow control and fluid dynamics ................................................................ 104
2.7 Photobioreactor preparation for use .................................................................... 105
2.8 Photobioreactor testing ........................................................................................ 106
2.8.1 Effect of flashing light on Chlorella emersonii growth ............................... 106
2.8.2 Preliminary testing ....................................................................................... 107
2.8.3 Photobioreactor and algal growth issues ...................................................... 109
2.9 Alternative airlift vertical reactor for testing of LED illumination; Chlorella
emersonii growth dependence on wavelength ............................................................... 117
2.9.1 Effect of lightconcentration on lipid profile and amount ............................ 124

2.10 Summary ............................................................................................................. 126
2.11 References ........................................................................................................... 130

3 Effect of environmental factors on fatty acid methyl esters composition in Chlorella
spp. 133
3.1 Lipid extraction ................................................................................................... 133
3.2 NMR spectroscopy initial screening ................................................................... 134
3.3 Fatty acid methyl ester profile determination using GCMS ................................ 136
3.4 Effect of carbon dioxide concentration on algal growth ..................................... 138
3.5 Design of experiments to study the effects of nitrate, phosphates and iron levels
and time on lipid profile and algal growth ..................................................................... 139
3.5.1 Growth of inoculation culture for design of experiments ............................ 142
3.5.2 Effect of nutrients and time on oleic acid methyl ester (C18(1)) synthesis . 143
3.5.3 Effect of nutrients and time on C16(4) synthesis ......................................... 148
3.5.4 Effect of nutrients and time on monounsaturated fatty acid methyl ester
production .................................................................................................................. 153
3.5.5 Effect of nutrients and time on polyunsaturated fatty acid methyl ester
production .................................................................................................................. 154
3.5.6 Effect of nutrients and time on saturated fatty acid methyl ester production157
3.6 Effect of nutrients and time on total lipid ............................................................ 160
3.7 Effect of time on algae growth and fatty acid methyl ester profile ..................... 163
3.8 Effect of carbon sources on algae growth and fatty acid methyl ester profile .... 166
3.9 Microwave extraction .......................................................................................... 170
3.10 Summary ............................................................................................................. 177
3.11 References ........................................................................................................... 180

4 Experimental .............................................................................................................. 183
4.1 General procedures .............................................................................................. 183
4.2 Lipid extraction ................................................................................................... 183
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4.3 Transesterification of lipids ................................................................................. 183
4.4 Nuclear magnetic resonance analysis .................................................................. 183
4.5 Mass spectrometry ............................................................................................... 185
4.6 Gas chromatography mass spectrometry (GCMS) .............................................. 185
4.7 Microwave technique .......................................................................................... 189
4.8 Acid value ............................................................................................................ 190
4.9 Bold’s Basal growth medium .............................................................................. 190
4.10 Cell counting ....................................................................................................... 190
4.11 Inoculation ........................................................................................................... 191
4.12 Design of experiments – effect of nutrients and time on algal growth ............... 192
4.13 Experimental design for mixotrophic study ........................................................ 194
4.14 Experimental design for toxicity studies ............................................................. 194
4.15 References ........................................................................................................... 196

5 Further work ............................................................................................................... 197

Appendix ............................................................................................................................ 198

vi
1 Introduction .................................................................................................................... 1
1.1 Preamble .................................................................................................................. 1
1.2 Biodiesel .................................................................................................................. 1
1.2.1 The requirement for biofuels as a transitional technology ............................... 2
1.2.2 Fatty acid methyl ester (FAME) profiles and fuel properties of biodiesel ...... 3
1.2.3 Issues with first generation biofuels ................................................................. 5
1.2.4 Potential of algal biofuels................................................................................. 7
1.2.5 Problems with algal biofuels .......................................................................... 10
1.2.6 Commercial ventures in algal biofuels ........................................................... 11
1.2.7 Policies and government goals ....................................................................... 11
1.2.8 Economical cost of algal biofuel .................................................................... 12
1.2.9 Energy costs and considerations .................................................................... 14
1.2.10 Transesterification .......................................................................................... 15
1.3 Microalgae ............................................................................................................. 17
1.3.1 Microalgal species .......................................................................................... 17
1.3.2 Nutrients ......................................................................................................... 19
1.3.3 Growth parameters ......................................................................................... 20
1.3.4 Growth rates ................................................................................................... 20
1.3.5 Algae growth conditions ................................................................................ 21
1.3.6 Effect of carbon sources on algal growth ....................................................... 22
1.3.7 Algal growth: the interrelated effects of carbon dioxideand pH ................... 22

1.3.8 Mixotrophic algae growth .............................................................................. 24
1.3.9 Salinity effects on algae growth ..................................................................... 25
1.3.10 Effect of limiting conditions on lipid content of algae .................................. 25
1.3.11 α-tocopherol and carotene .............................................................................. 29
1.4 Downstream processing and extraction of lipids from algae ................................ 30
1.4.1 Separation of algae from growth media ......................................................... 30
1.4.2 Lipid extraction utilising solvents .................................................................. 32
1.4.3 Alternative extraction techniques ................................................................... 33
1.4.4 Fatty acid methyl ester profile analysis .......................................................... 36
1.5 Photobioreactors .................................................................................................... 37
1.5.1 Considerations for photobioreactor design .................................................... 37
1.5.2 Algal reactor types ......................................................................................... 40
1.5.3 Examples of photobioreactors ........................................................................ 42
1.5.4 Mixing ............................................................................................................ 43
1.5.5 Carbon dioxide supply and control ................................................................ 45
1.5.6 Oxygen saturation .......................................................................................... 46
1.5.7 Mass flow control meter................................................................................. 47
1.5.8 Cleaning ......................................................................................................... 47
1.5.9 Light requirement and light emitting diodes .................................................. 47
1.5.10 Photosynthesis ................................................................................................ 49
1.5.11 Effect of light on algae growth and productivity ........................................... 52
1.5.12 Photoinhibition in photosynthesis .................................................................. 55
1.5.13 Light regimen effects ..................................................................................... 56
1.5.14 Effect of pulsed light on algal growth and acclimatisation ............................ 56
1.5.15 Effect of wavelength on algae growth ........................................................... 62
1.5.16 Carbon dioxide capacity and affinity in photosynthesis ................................ 63
1.6 Summary and aims ................................................................................................ 64
1.7 References ............................................................................................................. 66
3
1 Introduction
1.1 Preamble
Environmental change, such as carbon dioxide level increases, and global warming in
addition to current difficulties in fuel supply demand a cohesive and twenty-first century
solution. The need for alternative fuel sources has been well publicised by media and
science. Biodiesel should be at the forefront of the switch over to green energy as it
requires little, if any, modification of diesel engines as well as requiring minimal
integration into the fuel supply network. Algae are an excellent candidate for biodiesel
production due to their high bio-productive rate and the economic potential for valuable
product harvesting. Algae have also been proven to sequester carbon dioxide and shown to
reduce heavy metal contamination in waste water.1 The triglycerides present in algae can
be extracted and transesterified into biodiesel for use as liquid transportation fuel.2

This introduction will review relevant aspects of biodiesel production, algae cultivation
and photobioreactor design including the economical, energy and environmental outlooks
for algal biofuel.
1.2 Biodiesel
Biodiesel is the transesterification product, fatty acid alkyl ester, of triglycerides from land
crop oils, animal fats and more recently algal oils. As shown in Figure 1:1 the
transesterification reaction requires an alcohol and a catalyst (see section 1.2.10).3
ccaattaallyysstt
33 '' 33
TTrriiggllyycceerriiddee AAllccoohhooll FFaattttyy aacciidd aallkkyyll eesstteerr GGllyycceerrooll
((BBiiooddiieesseell))

Figure 1:1 – Transesterification reaction overview

1
1.2.1 The requirement for biofuels as a transitional technology
Two thirds of the whole world’s total energy consumption remains sourced from
traditional fossil fuels; hence the urgency due to diminishing fossil fuels to develop other
means of sustainable energy production.4 The increase in demand for petroleum,
particularly from emerging markets such as Asia, is increasing the requirement to expand
renewable fuels, specifically liquid fuels like biodiesel.2 The transportation industries,
including aviation and shipping rely heavily on liquid fuel derived from petroleum.5
Biofuels are needed to improve energy security (see section 1.2.7), climate change and
rural development.6 Other issues which biofuels can address are increasing global
population, mounting waste, depleting resources and increased environmental pollution.7

Biodiesel can have comparable energy content and similar physical properties, such as
cetane number and cold pour point, to petroleum diesel thus making it a desirable target for
biofuel production. The use of transesterified plant oil as a fuel is credited to E. Duffy and
J. Patrick in 1853, whilst R. Diesel showcased the first engine running from vegetable oil
in 1900 at the International Exhibition, Paris.8 The three main targets for any biofuel are to
have (i) high energy density with regards to mass and volume; (ii) to be producible at
yields around the stoichiometric maximum for the chosen biomass starting material; and
(iii) to be compatible with the existing fuel distribution infrastructure.9

Mass changeover to electrical and hydrogen powered terrestrial transportation is expected
to begin within the next couple of decades; this widespread uptake can then be more fully
developed.4-5 Biofuels, particularly biodiesel, can fit this energy gap and help ease
predominantly industrialised countries into a future without dependence on fossil fuels.
Biofuels are required to have beneficial properties, such as high density liquid energy, fuel
security and easily biodegradable products (see section 1.2.4).2, 10 It is sought-after that the
biofuel has a lower carbon footprint than that of the fossil fuel it replaces and that the
biofuel is suitable for the use it is being employed for. It may also be considered that
subsidies may be required for renewable biofuels to be commercially available. It is
desirable that the production of high value co-products alongside biofuel will allow biofuel,
especially biodiesel to become economically viable. However, when high value products
are created on the large scale required for fuel production they may lose their high-value
status due to market saturation.11
2