Anaerobic Degradation of Linoleic (Clsa), Oleic (C18:i - T-Space

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Anaerobic Degradation of Linoleic (Clsa),O leic (C18:i)a nd Stearic (Clsa) Acids and
their Inhibitory Effects on Acidogens, Acetogens and Methanogens
Jerald David Anthony Lalman
A thesis submitted in conformity with the requirements
For the degree of Doctor of Philosophy
Graduate Department of Civil Engineering
University of Toronto
OCopyright 2000 by Jerald Anthony David Lalrnan
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Anaerobic Degradation of Linoleic (Cl82), Oleie (Ci8,i)a nd Stearic (Cis:o) Acids and
their Inhibitory Effects on Acidogens, Acetogens and Methanogens
Jerald David Anthony Lalman
Doctor of Philosophy
Department of Civil Engineering
University of Toronto
2000
Abstract
Effiuents fiom many food processing industries contain fats and oils in addition to
carbohydrates and proteins. Long chain fatty acids (LCFAs), a hydrolysis byproduct of
fats and oils, are difticult to degrade and are inhibitory to anaerobic organisms. These
acids are degraded via f3-oxidation but the compound initiating the mechanism has not
been clearly identified. Although LCFAs inhibit aceticlastic methanogenesis their effects
on acidogenesis, acetogenesis and hydrogenotrophic methanogenesis have not been well
studied.
This study assessed the degradability of linoieic (C oleic (Ci and steark (C
8:?), 1)
acids and determined their inhibitory effects on anaerobic organisms in 160 mL s e m
bottles. Degradation and inhibition studies were conducted using 10, 30, 50 and 100
rng~-L'C FA. Inhibition studies using glucose, butyrate and acetate (each at 100 rng-~-')
and hydrogen (10.1 kPa) investigated the effects of the three LCFAs on acidogens,
acetogens, acet iclastic methanogens and hydrogenotrophic methanogens.
Unsaturated C 18 LCFAs were degraded to shorter chah LCFAs however, no LCFA
byproducts were detected as intermediates during the degradation of stearic (C18:o)a cid.
Palmitic and myristic (CI~:oa)c ids were produced fiom linoleic CI^:^) acid at al1
concentrations examined and in cultures receiving more than 10 rng-~-oll eic (Ci8:la) cid.
In cultures receiving 100 rng-~''L inoleic (C 1 a2) acid both oleic (C 1 !) and palrnitoleic
(C 16:1)a cids were detected.
Acidogenesis was af5ected by the presence of LCFAs and synergistic inhibitory
effects of al1 three acids on acetogenesis were observed. Hydrogenotrophic
methanogenic inhibition was observed and aceticlastic methanogens were inhibited at al1
LCFA concentrations examined. in cultures fed with linoleic (C 1 g:2) and oleic (C 18: I)
acids, inhibition of acetate methanogenesis was concentration dependent but for cultures
receiving stearic (C ls:o)a cid, the effect was independent of concentration.
In comparison to stearic CI^:^) acid, iinoleic (Ci8:2)a nd oieic (Cla:~a)c ids were
degraded faster. Therefore, the design of a full-scaie system will depend on the SRT for
the more slowly degrading LCFA. LCFAs affected glucose and butyrate degradation.
Hence, in effluents containing carbohydrate and LCFAs mixtures, the degradation of
carbohydrate monomers wil be afTected. In comparison to oieic (Cls:~a)c id, greater
aceticlastic inhibition was observed for cultures receiving linoleic (Clg:2) acid. Thus, it is
recornmended a two-stage process, acetogenic followed by rnethanogenic, be used to
minimize the inhibition.
...
III
Acknowledgement s
This w-ork is dedicated to my mother. Rosaline Lalman, who passed away while
n~itingth is thesis. My father. Samuel Lalman. has been a great inspiration throughout
my life and 1 n-ish to thank him for instilling into me the meaning of accomplishrnent. 1
would also like to give my special appreciation to Nin for her precious suggestions. her
stronp encouragement at times of difficulty. love and understanding throughout this
u-hole process. Also. thanks to my brother. Edmund Lalman and my sister. Maria
Crutchley for sharing these troubled times in our lives.
I wish to espress my sincere appreciation to my advisor. Dr. David Bagley for his
intelligent supervision. constructive criticisms. inspiration and friendship. My sincere
appreciation estrnds to m). other cornmittee members Dr. Grant Allen. Dr. Don Kirk and
Dr. Brrnr Slecp. n-hose guidance. assistance and friendship are also invaluable. 1 am also
~ratrsfulto Dr. Mary Jans Philips and Durga Prasad for their encouragement and ad~vicr.
C
1 n.ould like to th& Rajesh Seth. Russell D'Souza and Yale Zheng for sharing their
kno\vledgs and helpful comments.
Financial assistance was providsd by the University of Toronto and the Ontario
Ministr).. of Energy Science and Technology: Singapore-Ontario joint research
programme.
Finall>-.1 would like to t h d t he Department of Civil Engineering for their support
during the four and one-half years of this study.
Table of Contents
Page No.
..
Abstract 11
Acknowledgements
List of Tables 5
List of Fi,w res sii
1.1 Context
1.2 Research Objectives
1.4 Publications
2.2 Fundamentals of Anaerobic Wastewater Treatment
2.2.1 Hydrolysis
2 -2.2 Acidogenesis
2.3.3 Acetogenesis
2.2.3 Methanogenesis
2.3 Anaerobic Reactor Technologies Used to Treat Emuents
Containing Long Chain Fatty Acids
2.3.1 Introduction
2.3 -2 LOWR ate Treatment
2.3.3 High Rate Treatment
2.4 Long Chain Fatty Acids
2.4.1 Sources and Treatment
2.4.2 Composition and Structure
2.4.3 Biodegradation of LCFAs
2.4.4 Effects of Hydrogen and Volatile Fatty Acid
A. Hydroger~
B. 1,klafileF atp Acidr
2.4.5 Inhibitory Effects
A. Effecrso n Membrane Fmcîion
B. Effecrso n Anaerobic Orgmrisms
2.4.6 Factors AfFecting LCFA Degradation and Inhibitory Eftècts
A. Szi bstra~eM olemlar Sb-uctlrre and Conceritratiorr
B. Tentperarure Eflects
C. Soltrbilzty Efects
D. pH EfJecrs
E. Coslibsirare ami fiermocjtraniic Ef/ecrs
Summary
MATEMALS AND METEODS
Experimental Plan
Reagents
Batch Reactors
3.3.1 Inoculum Source
3.3.2 Operation of Inoculum Reactors
Hydrogen and Methane Measurements
Volatile Fatty Acid (VFA) Measurement
Development of a LCFA Delivery Strategy
Long Chain Fatty Acid (LCFA)M easurement
3.7.1 LCFA Extraction- Method Development
3.7.2 LCFA Extraction- Phase Partitioning Studies
3.7.3 LCFA .4naiysis
Serum Bottle Preparation
Glucose Measurement
Total Suspended Solids (TSS), Volatile Suspended Solids
(VSS), Alkalinity and pH hleasurements
BATCH REACTORS
Ex~erimentalR esults
4.2 Discussion of Results 84
DEGRADATION OF LINOLEIC CI^:^) ACID AND ITS
INHIBITORY EFFECTS ON METHANOGENESIS
Experimental Results
5.1 -1 Linoleic Acid Degradation
5.1.2 Inhibitory Effects of Linoleic (C Acid on
Methanogenesis
A. Acetate Degradatiorz
B. Hwogen Cor~sumptisrt
i. Hydrogen uptake 1 day after linoleic (C18:2)a cid
addition
ii. Hydrogen uptake 18 days and 35 days after
linoleic (C acid addition
18.~)
Discussion of Results
5.2.1 Linoleic Acid Degradation
5 -2.2 Linoleic Acid-Methanogenic Inhibition Studies
Summary 104
DEGRADATION OF OLEIC (Cl~t:A~C) ID AND ITS INHIBITORY
EFFECTS ON METEIANOGENESIS 105
Experimental Results 10s
6.1 .1 Oleic (Cls:l)A cid Degradation 105
6.1.2 Inhibitory Effects of Oleic (Cls:l)A cid on Methanogenesis 1 08
A. A ce tare Degrndatiort 1 08
B. Hydrogen Corrs~rmptior~ 110
Discussion of Results
6.2.1 Oleic (C18:lA) cid Degradation
6.2.2 Oleic (C Acid-Methanogenic Inhibition Studies
18:1)
Summary 115
DEGRADATION OF STEARIC (Cls:o) ACCD AND
ITS iNHIBI TORY EFFECTS ON METHANOGENESIS
Experimental Results
7.1.1 Stearic (Cls:o)A cid Degradation
7.1.2 Inhibitory Effects of Stearic (Cis:,) Acid on
Methanogenesis
A. Acetate Degradation
B. Hydrogen Consrrmptio?l
7.2 Discussion of Results
7.2.1 Stearic (C18:0) Acid Degradation
7.2.2 Stearic (C ls:o) Acid-Methanogenic Inhibition Studies
7.3 Summary
8.0 COMPARISON OF LCFA DEGRADATION STUDIES
8.1. Products of Linoleic CI^:^), Oleic and Stearic (C18:o) Acids
Degradation
8.2 Possible Pathways for Linoleic (Cls:i) Acid Degradation
9.0 INBIBITORY EFFECTS OF LINOLEIC (Cls:r),O LEIC (Cis:,)
AND STEARIC (Cls:o)A CIDS ON ACIDOGEKIC, ACETOGENIC
AND METHANOGENIC ACTWITI'
9.1 Experimental Results
9.1.1 Acidogenic Inhibitory Effects-Glucose Degradation
9.1 .2 -4cetogenic Inhib itory Effects-Butyrate Degradation
9. 1.3 Methanogenic Lnhib itory Effects-Hydrogen Consumption
9.2 Discussion of Results
9.2.1 Glucose Degradation
9.2.2 Butyrate Fermentation
9.2.3 Hydrogen Consumption
9.3 Summary
10.0 SUIM31ARY OF METHANOGENIC STUDIES AND
LCFA INHLBITION MECBANISM
10.1 Methanogenic Studies
10.1. 1 Aceticlastic Methanogens
10.1 .1 Hydrogenotrophic Methanogens
10.2 LCFA Inhibition Mechanism
11.0 SUMlMARY AND CONCLUSIONS
11.1 Summary
1 1.1.1 LCFA Degradation
1 1 -12 LCFA Inhibition
A. A cidogem
B. Acefogens
C. Methmiogerzs
i. Aceticlastic methanogens
ii. Hydrogenotrophic methanogens
1 1.2 Conclusions
12.0 ENGINEERING SIGNIFKANCE AND SUGGESTIONS
FOR FUTURE RESEARCH
13.0 REFERENCES
Appendix A: LCFA Free Energy Cakulations
Appendix B: Example of LCFA Mass Balance Calculations