preparation of volatile fatty acid (vfa) calcium salts by anaerobic digestion of glucose

This article has been accepted for publication and undergone full peer review but has not been through the copyediting,

typesetting, pagination and proofreading process, which may lead to differences between this version and the Version of

Record. Please cite this article as doi: 10.1002/bab.1301.

This article is protected by copyright. All rights reserved. 1

Preparation of volatile fatty acid (VFA) calcium salts by anaerobic digestion of glucose

Xiaofen Li*, Janis E. Swan, Giridhar R. Nair and Alan G. Langdon

School of Engineering, The University of Waikato, Private Bag 3105, Hamilton, New Zealand 3240

Abstract

Many potentially useful intermediates such as hydrogen and volatile fatty acids (VFAs) are formed during the

complex anaerobic digestion processes that produce methane from biomass. This study recovers VFAs from an

anaerobic digester by a combination of gas stripping and absorption with calcium carbonate slurry. Glucose was

used as the model substrate because it is readily available, inexpensive and easily digested. Sludge from a meat

works anaerobic digester produced methane and carbon dioxide (and sometimes a small amount of hydrogen)

when batch-fed with glucose. Conditioning the neutral anaerobic sludge to an acidic pH (below 4.8) was

achieved using repeated 1-g L-1

doses of glucose. After conditioning, mainly VFAs and hydrogen were

produced. The intermediate VFAs could be stripped using headspace gas. In subsequent fed-batch

digestion/stripping cycles, the pH decreased when glucose was added and then rose when the VFA was gas

stripped. The predominant acids formed at low pH values were lactic, butyric and acetic acids. Lactic acid was

converted to VFAs during stripping. The VFA calcium salts recovered were 80% butyrate, and 20% acetate with

minor quantities of propionate and valerate.

Keywords:

Anaerobic digestion, bioreactors, volatile fatty acid, stripping, absorption, recovery

1. Introduction

The search for sustainable and environmentally-compatible alternatives to fossil fuels for producing organic

commodity chemicals has led to renewed interest in converting biomass using microbial processes. Anaerobic

digestion is a multi-step series of hydrolytic, acidogenic, acetogenic and methanogenic biological reactions that

converts complex polymeric substances to simpler molecules [1]. This process is routinely used to process

waste biomass from food processing, agriculture and municipal wastewater treatment into stabilized sludge, a

nutrient-rich effluent, methane and carbon dioxide. Hydrogen and volatile fatty acids (VFAs) are formed as

intermediates. These are converted to methane in the final methanogenic step. Hydrogen and VFAs have

higher values as industrial chemicals than methane [2], which provides an incentive to find economic methods

for their recovery.

Although there have been many reports on controlling anaerobic digestion to produce hydrogen [3-9], there are

fewer reports on recovering VFAs. A process for removing VFAs could have the added benefit of preventing

the “souring” problem when trying to maintain continuous anaerobic fermentations [10]. The purpose of this

research was to condition an anaerobic sludge to an acidic pH to inhibit methanogenesis and ensure that VFAs

were predominantly in their volatile undissociated forms, and then to apply a recently reported procedure [2, 10]

to strip and recover VFAs as VFA calcium salts.


1.1 Volatile fatty acid stripping and recovery

Continuous operation of an anaerobic digester to produce acetic acid and other VFAs involves inhibiting

methanogenesis and simultaneously removing and recovering the VFAs. Literature indicates that liquid/liquid

extraction and reactive extraction using tertiary amines [11] can be used to recover acetic acid from dilute

solutions. These methods are not compatible with maintaining an active fermentation. Recently, a more

suitable gas stripping method, based on volatility of the VFAs, has been developed [2, 10]. While gas stripping

is routinely used to remove volatile contaminant components from liquid process streams, there is little literature

on using this process to remove and/or recover VFAs from dilute solutions of processing waste streams or

anaerobic digesters. This may due to VFAs having much higher Henry’s law constant values compared with

strippable components such as hydrogen sulphide, ammonia, carbon dioxide, etc [12]. Conventional stripping is

successful when the volatile solutes have Henry’s Law constants less than 10 mol L-1

atm-1

[13]. Values for

VFAs are more than two orders of magnitude higher than this and the concentrations of undissociated volatile

forms of VFAs are pH dependent (the higher the pH, the lower the concentration).

Gas stripping used in industrial processes such as wastewater treatment needs to be rapid and as near complete

as possible. This is favored if Henry’s law constants are small. However, because anaerobic digestion is a slow

process, even slow gas stripping of VFAs, can be effective. An optimized continuous process only requires that

stripping remove the acids as fast as they are produced. The stripping should maintain the pH at a level that

avoids excessive VFA dissociation and inhibition of the digestion.

1.2 Inhibiting methanogenesis

To obtain high VFA concentration, methanogenesis, which converts VFAs to methane, must be inhibited. This

can be achieved by acidification [14, 15, 16], exposure to air [6, 17], thermal treatment [4, 6, 7, 18] or chemical

inhibition with, for example, halogenated hydrocarbons [8]. Lowering the pH will also satisfy the requirement

that the VFAs are in their volatile, undissociated forms and therefore strippable. In conventional anaerobic

digestion, the aim is to maintain a neutral pH. Methane production falls markedly when pH is below 6.3 or

above 7.8 [15, 19]. Lowering bioreactor pH directly by adding acid or indirectly by overloading with easily-

digested substrate are possible ways of inhibiting activity of methanogens and favoring the acidogens and

acetogens that produce hydrogen and VFAs. Panamas et al. reported that consumption of large amounts of

glucose lowered pH and produced high VFAs concentrations, however, glucose concentrations of 10 g L-1

and

greater are toxic for the anaerobic microbes [20]. When methanogenesis has been suppressed, acid

accumulation during batch fermentation will eventually lower pH to levels that inhibit further fermentation and

the reactor “sours”. Hydrogen (and lack of methane) in the gaseous products can be correlated with

accumulation of VFAs in the bioreactor fluids [9], allowing gas composition to indicate VFAs accumulation, at

least while the reactor is producing gas.

2. Materials and methods

2.1 Fed-batch bioreactor/stripping/recovery system

The recently reported technology for stripping VFAs from aqueous systems [10, 12] was applied to a bench-

scale reactor (Figure 1) made of a 5-L Pyrex bottle bioreactor containing 1.7 L sludge. The 1-L conical flask

recovery system contained 200 mL 2M CaCO3 slurry. The bioreactor contents, containing dilute VFAs, were

stripped using water-saturated nitrogen gas entering via a 7-mm (i.d.) open tube. The gas containing the VFAs

stripped from the bioreactor was then passed through a 19-mm (i.d.) open tube immersed in the CaCO3 recovery


system. The peristaltic pump was fitted with 166 cm silicon tube (6.4 mm i.d., Masterflex 6404-17) and set to

deliver 1.06 L min-1

(stripping rate of 0.62 L-1

L-1

min-1

). The system was maintained at 35oC in a modified

incubator.

The VFAs of interest (acetic, propionic, butyric, and valeric) all have pKa values close to 4.8. All have similar

Henry’s law constants [12] (1.1 -5.7) x 103. Henry’s Law describes the partial pressure of a volatile component

(e.g. VFA) in a gas phase (PVFA) in equilibrium with a dilute solution of that component at a concentration,

CVFA(aq):

VFA(aq) H VFAC = K P (1)

where, KH is the Henry’s Law constant (mol L-1

atm-1

).

The maximum transfer rate (TR) of a specific VFA that can achieved at a stripping gas volumetric flow rate, Q,

is:

VFA(aq)

VFA(g)

H

CTR= Q C = Q x

K RT

(2)

Existence of this relationship between TR and Q will demonstrate VFA equilibration with the gas stream during

stripping and will also indicate that VFA is quantitatively removed from the gas stream in the recovery system.

Once the VFAs in the gas stream have been transferred into the CaCO3 recovery system, the gas can absorb

more acid when it is recirculated through the bioreactor. The stripping gas remains water saturated and at

constant temperature.

Previous research [10] showed that Equation 2 was valid for nitrogen flow rates up to 2.0 L min-1

when

stripping acetic acid from dilute solution and then recovering the acetic acid in CaCO3 slurry, indicating that

liquid-vapor equilibrium was established during stripping and quantitative removal of the stripped acid was

achieved during the recovery step.

2.2 Anaerobic sludge

Anaerobic sludge samples were obtained from the ambient temperature plug flow reactor at a local meat

processing company. The total solids (TS) of the sludge was 63 g L-1

. Of this, 59% (37 g L-1

) was volatile

solids (VS). As-collected sludge alkalinity, 5 day carbonaceous biochemical oxygen demand (BOD5), chemical

oxygen demand (COD), ammonia (NH4) nitrogen as N, total suspended solids (TSS ), total phosphorus as P

were 2008 mg CaCO3 L-1

; 758 mg L-1

, 944 mg L-1

, 457 mg L-1

, 390 mg L-1

and 67 mg L-1

respectively. The

high ammonium and phosphorous content were considered to be sufficient to maintain microbial activity

without further additions. Anecdotal evidence indicated this sludge often produced hydrogen and hence might

be readily conditioned to produce VFAs. When operated at 20oC and batch-fed with a single 1 g L

-1 dose of

glucose, the sludge produced a biogas containing 2.7% hydrogen. The sludge stored at 20oC and fed

periodically with glucose but no other nutrients retained good activity during the trials.

Insert Fig. 1 here

2.3 Fed-batch conditioning experiment (Cycle 1)

Anaerobic sludge (1.7 L at 6.3% volatile solids) was transferred to the 5-L bioreactor and conditioned to acid

pH by adding 1-g L-1

doses of glucose at the loading rate of 0.05 g glucose (g VS)-1

. Each successive addition


was made after gas production and pH lowering from the preceding dose had ceased (usually after 1 to 3 days).

After each glucose dose, the system was flushed with oxygen-free nitrogen to remove any introduced oxygen.

Ambient temperature (20oC) was maintained during the conditioning process because the sludge had been

collected and stored at ambient temperature.

Gas composition during digestion was monitored regularly by in-line gas chromatography (GC). The gas

produced was collected over water (pH 7.32 ± 0.05) in inverted 250, 1000 or 2000-mL measuring cylinders.

The pH in the measuring cylinder water did not change, indicating negligible adsorption of carbon dioxide. The

batch feeding was continued until the reactor pH was lowered to pH 4.65

2.4 Fed-batch digestion /stripping/recovery cycles

After conditioning of the bioreactor sludge at 20oC (with 15 1-g L

-1 additions of glucose), the reactor contents

were gas stripped with head space gas (mainly nitrogen) at 35oC until pH increased to approximately pH 5.0

(Cycle 1). Three further successive fed-batch digestion /stripping/recovery cycles were performed, all at 35oC

(Cycles 2, 3 and 4). Stripping was started after the pH in the reactor had dropped below pH 4.8. The glucose

digestion step of Cycle 1 was the 15 successive 1-g L-1

doses at a loading rate of 0.046 g glucose (g VS)-1

needed to condition the sludge to acid pH. In Cycle 2, six successive glucose additions were batch-fed as 1-g L-

1 doses. In Cycle 3, glucose was added as nine 1-g L

-1 doses at a loading rate of 0.046 g glucose (g VS)

-1,

followed by one 5-g L-1

dose at loading rate of 0.23 g glucose (g VS)-1

and then one 10-g L-1

glucose dose at

loading rate of 0.46 g glucose (g VS)-1

. In Cycle 4, in which there was no stripping step, one 10-g L-1

dose was

added at a loading rate of 0.46 g glucose (g VS)-1

. After the initial conditioning at 20oC, the reactor was run at

35oC. Total glucose added during each cycle was 15 g L

-1, 6 g L

-1, 24 g L

-1 and 10 g L

-1 for cycles 1, 2, 3 and 4

respectively. Individual additions during each cycle were kept at 10 g L-1

or less to avoid the possibility of

glucose toxicity. Generally, digestion of the successive additions was complete within a few days. In cycle one,

the 15 glucose additions were added over a period of 79 days allowing approximately 5 days for digestion after

each addition. For cycles 2 and 3, the average digestion times were 1.7 and 4.4 days. Stripping was continued

for 9, 10 and 67 days for cycles 1, 2 and 3 respectively. For cycle 2 the stripping time was kept approximately

the same as the total digestion time to test the possibility of operating the reaction under conditions of

continuous simultaneous digestion and stripping.

2.5 Gas analysis

Head space gas samples were analyzed using a Perkin-ElmerTM GC with a thermal conductivity detector

(TCD) and a column sequence of HaySep Q (80/100), molecular sieve 13X (pore size 13 Å with sodium as the

primary cation) and HaySep D (100/120) columns. The molecular sieve separated hydrogen, oxygen, nitrogen,

methane, and carbon monoxide. Because carbon dioxide is irreversibly adsorbed by the zeolite, the GC-TCD

program was set to reverse gas flow before carbon dioxide had passed through the HaySep Q columns. The

following program was used: oven temperature 50oC, detector temperature 250

oC, inject temperature 120

oC,

carrier gas argon (90 psi.), flow rate 20 mL min-1

, program time 10 min with reversed flow at 2.65 min. Gas

flow was reversed using pneumatic switches operated with dry air at 65 psi.

The GC system was calibrated with a standard gas mixture (Matheson Tri. Gas, Grace Davison Discovery

Science, USA) at regular intervals during the trials. Peak areas had coefficients of variation (CVs) of 2.5%,

2.7%, 2.3%, and 5.0% for H2, CH4, N2 and CO2 respectively. Head space gas samples from the bioreactors were

injected at room temperature to the columns via a 1-mL sample loop operated pneumatically by a dry air supply.


2.6 Analysis of digester contents

Digester contents (glucose, lactic acid, acetic acid, propionic acid, iso-butyric acid, butyric acid and valeric acid)

were analyzed by HPLC [21]. Samples of reactor fluid (1 mL) taken at regular intervals during the runs were

stored at -20oC until analyzed. When required, samples were centrifuged at ambient temperature and 12,000

rpm for 10 min during which the samples thawed and produced a clear supernatant. A 1-mL sample of the

supernatant was withdrawn using a 1-mL syringe fitted with a 22 G x 1 ½ (0.71 x 38 mm) sterile single use

needle and filtered through a 0.20-µm minisart®

-plus syringe filter. Forty µL of 7% sulfuric acid was added to

the filtered supernatant to convert the weak acids to their protonated forms and help maintain functionality of

the column.

The samples were shaken in a vortex (MS1 Minishaker IKA® Works, Malaysia) mixer for about 10 seconds and

then analyzed by HPLC (Waters 515 Pump, Waters Column Heater Module, Waters 996 Photodiode Array

Detector (PDA), Waters 2414 Refractive Index (RI) Detector and a Rheodyne 7725i manual sample injector

(20-µL sample loop)). The system was controlled using Waters EmpowerTM

2 Chromatography Software. The

analyses were performed isocratically at 0.6 mL min-1

and 60oC with a 300 x 7.8-mm i.d. cation exchange

column (Aminex HPX-87H) equipped with a cation H+ microguard cartridge (Bio-Rad Laboratories). Mobile

phase was 5 mmol L-1

sulfuric acid prepared with milliQ water, filtered through a 0.2-µm minisart®-plus

syringe filter and degassed under vacuum in an ultrasonic bath for 20 min. Peak areas were highly reproducible

with CVs of 0.54%, 0.23%, 1.26%, 1.39%, 3.45%, 3.17% and 1.05% for the acetic acid, glucose, propionic acid,

iso-butyric acid, butyric acid, valeric acid and L-lactic acid standards respectively.

3. Results and discussion

3.1 Conditioning the sludge with glucose substrate

Gas production at 20oC (collected over water) for 1 g L

-1 glucose doses leveled off to approximately 190 mL (g

glucose)-1

. After each successive glucose dose had been digested, pH decreased. Initially the pH drops were

about 0.1 pH units but increased to about 0.3-0.4 pH units below pH 6.0 and then by 0.1 pH units at pH below

5.0. After 15 doses, pH had decreased from 7.5 to 4.65. The sludge that had been producing predominantly

methane and carbon dioxide was conditioned to one that produced hydrogen and carbon dioxide and no

methane.

3.2 Fed-batch digestion and gas stripping cycles

Compositions of the reactor fluid were determined before digestion (BD), after digestion (AD) and after

stripping (AS) for three complete fed-batch/stripping /recovery cycles (Cycles 1, 2 and 3). The BD data for

Cycle 1 is for the unconditioned sludge. The AS and BD data for the successive batch fed/stripping cycles are

similar. The results (Table 1) show that glucose is converted to predominantly butyric acid, acetic acid and

lactic acid, which is consistent with the pathways for anaerobic digestion of glucose. At neutral pH, glucose is

converted predominantly to methane and carbon dioxide. As pH decreases, methane production decreases,

hydrogen production increases and VFAs (mainly acetic acid and butyric acid with smaller amounts of lactic

acid) accumulate. These transformations, which depend on the conditions, can be described by the overall

stoichiometries [23, 24, 25].

6 12 6 3C H O 3CH COOH

6 12 6 3 2 2 2 2C H O CH CH CH COOH + 2CO + 2H


6 12 6 3C H O 2CH CHOHCOOH

The predominance of butyric acid rather than acetic acid in the supernatant is consistent with the presence of

significant volumes of hydrogen in the head space gas. The stoichiometries and Gibbs free energy changes for

forming acetic acid from the higher carbon VFAs [26, 27] mean there will little tendency for any butyric acid

formed to be converted to acetic acid when hydrogen is present.

- - +

3 3 2 2 3 2CH CH CH COO + 2H O 2CH COO +2H +2H , ΔGo' = 48.3 kJ mol

-1

- - + -

3 2 2 3 3 2CH CH COO + 3H O CH COO +H +HCO +3H , ΔGo' = 76.1 kJ mol

-1

- -

3 2 2 3 2 2CH CH COO + 2H O CH COO +3H +CO , ΔGo' = 71.7 kJ mol

-1

During stripping, pH always increased and lactic and butyric acid concentrations always decreased but there was

no consistent trend for acetic acid (Table 2). No VFA was completely stripped. The disappearance of the non-

volatile lactic acid during acidic anaerobic stripping indicates its conversion to VFAs, as reported previously

[22]. Acetic and propionic acid contents in the supernatant did not appear to be significantly affected by

stripping. The acid stripped at low pH values appeared to be replaced by acids formed in their unstrippable

anionic forms at the higher pH values towards the end of stripping. It is known that acetate-producing microbes

become active as pH rises, allowing for head space H2 and CO2 to be converted to acetate. Alternatively, if

lactic acid is converted to acetate during the higher pH stages of stripping, this acetate would also remain largely

unstripped. The decrease in HAc after stripping during cycle 3 but its increase during cycles 1 and 2 can be

explained by the fact that the pH during the stripping of cycle 3 remains below pH 4.95. The lower pH would

allow HAc to be more exhaustively stripped while inhibiting the methanogenic processes that would allow un-

strippable acetate to be formed from higher C acids and CO2 and H2.

Data for pH changes during the digestion and stripping steps are summarized in Table 2. Changes in pH should

relate to changes in concentrations of the acid and salt species present. In the case of a single weak acid and its

salt, the relationship between acid and salt concentrations is the well-known Henderson equation [26]:

a 10

[A ]pH = pK + log

[HA]

Insert Table 1 here

While it is possible to devise individual equations for systems of mixed acids and their salts, because the pKa

values of the VFAs are all close to 4.8, an approximate estimate of pH can be made by summing the

concentrations of all the undissociated acids and their salts and using a pKa of 4.80 (the mean pKa value of the

two main VFAs present, butyric acid and acetic acid). The total VFA salt concentration in the system will

depend on the initial alkalinity of the sludge and will remain essentially constant. It was estimated from half the

total VFA content (salt and undissociated acid) of Cycle 2 after digestion where the pH of 4.78 (Table 1), was

very close to the mean pKa value so that salt and undissociated acid concentrations would be approximately the

same. The observed and calculated pH values for the various systems investigated (Table 2) were usually

within ± 0.1 pH units for all systems except those with significant concentrations of lactic acid (Cycle 1 AD,

Cycle 4 AD (1) and AD (2)). The presence of lactic acid (pKa 3.86) or other acid species such as polylactic acid

(pKa = 3.1) [29], which have pKa values lower than 4.80, will produce a lower pH for the given carboxyl


content. For example, the observed pH of 3.85 for Cycle 4 AD1 is close to the value calculated by considering

only the lactic acid equilibrium. The lower than calculated pH of 3.58, recorded after further standing for Cycle

4 AD2, indicates the presence of a species with a pKa even lower than that of lactic acid.

Insert Table 2 here

To determine carboxylic acid yields and carbon conversion efficiency during digestion and stripping, the

compositions of the supernatant and headspace gas were used to calculate moles carboxylic acid formed and

moles of carbon associated with each product species. Comparing the total moles of carbon identified as acid

and carbon dioxide products with moles of carbon added as glucose (Table 3) indicates that carbon recovery was

65%, 58% and 77% for cycles 1, 2 and 3 respectively. The glucose carbon not accounted for in these products

is likely to have been used in other digestion metabolites produced, including polymeric material detected (but

not quantified) as a significant peak at the column void volume and cellular biomass that would have been

removed from the samples during sample preparation. The fact that conversion efficiency to acids was greatest

for cycle 3 where pH was generally much lower than for the other cycles indicates that the formation of the

other metabolic products was inhibited by low pH. In general, yields for cycles 1, 2 and 3 were similar (cycles 1

and 3) or lower (cycle 1) than those reported in the literature for similar conditions [30] consistent with the

absence of contributions from breakdown of biomass during these experiments. However the anomalous

behaviour during cycle 4 is of interest. This system was not stripped because the lack of gas evolution after

adding glucose was initially interpreted as loss of bioactivity. After 23 months at room temperature, pH had

dropped to 3.58 but no gas had been produced. Total dry solids content decreased from 6.3 wt% to 4.3 wt% and

a carbon balanced indicated that supernatant lactic acid contained more carbon than could have been produced

from the added glucose (Table 3). It appears that some of this lactic acid had been produced from the dry solids

that had been consumed.

Insert Table 3 here

3.3 Recovery of crystalline product

In early work recovery of pure product was compromised by carry-over to the CaCO3 recovery system of foam

produced in the bioreactor. Foaming was eliminated by adding SE-15 anti-foaming agent (Sigma, USA). A

pure salt solution was then obtained as filtrate from the CaCO3 slurry. The XRD profile of the 3.46 g of white

crystalline product obtained from the evaporated filtrate collected from pH 4.8 to 4.95 for stripping cycle 3

indicated the presence of VFA salts but no calcium carbonate. However, the individual components could not

be identified further from XRD. The HPLC analysis of the white powder confirmed it consisted of calcium salts

of VFAs, predominantly butyric acid (Table 4).

Insert Table 4 here

4. Conclusions

Neutral methane-producing anaerobic sludge can be conditioned to a hydrogen-producing acidic sludge by

repeated dosing with glucose. At pH values below 4.8, VFAs are formed in their undissociated volatile form


and can be gas stripped from the reactor and recovered as calcium salts, predominantly calcium butyrate, by

passing the stripping gas through calcium carbonate slurry. Prolonged gas stripping can raise the pH of an

acidic digester to pH 5. The operation of an anaerobic digester under acidic conditions, using appropriate

substrate addition and gas stripping rates to remove VFAs as fast as they are formed, provides a possible new

method for continuous conversion and recovery of VFAs (particularly butyric acid) from suitable biomass

substrates. These might include biomass wastes of high sugar or starch content such as fruit and food

processing wastes.

5. Acknowledgment

The authors gratefully acknowledge the financial support of WaikatoLink Ltd and The University of Waikato,

Hamilton, New Zealand.

6. References

1. Evans, G, ed. (2001) Anaerobic digestion, in: Biowaste and Biological Waste Treatment, James &

James, London, 94-95

2. Langdon, A and L Li (2010) Anaerobic digestion: Harvesting high value by-products. NZ553786

3. Baghchehsaraee, B, Nakhla, G, Karamanev, D and Margaritis A (2010) Fermentative hydrogen

production by diverse microflora. Int J Hydrogen Energy 35:5021-5027

4. Fan, Y, Li C, Lay JJ, Hou H, and Zhang G (2004) Optimization of initial substrate and pH levels for

germination of sporing hydrogen-producing anaerobes in cow dung compost. Bioresour Technol

91:189-193

5. Lay, JJ (2001) Biohydrogen generation by mesophilic anaerobic fermentation of microcrystalline

cellulose. Biotechnol Bioeng 74: 280-287

6. Li, Z, Wang, H, Tang, ZX, Wang, XF and Bai JB (2008) Effects of pH value and substrate

concentration on hydrogen production from the anaerobic fermentation of glucose. Int J Hydrogen

Energy 33:7413-7418

7. Liang, DW, Shayegan SS, Ng, WJ and He, J (2010) Development and characteristics of rapidly formed

hydrogen-producing granules in an acidic anaerobic sequencing batch reactor (AnSBR). Biochem Eng

J 49:119-125

8. Wang, CC, Chang, CW, Chu, CP, Lee, DJ, Chang, BV, Liao, CS and Tay, JH (2003) Using filtrate of

waste biosolids to effectively produce bio-hydrogen by anaerobic fermentation. Water Res 37:2789-

2793

9. Zhu, H, Parker, W, Basnar, R, Proracki, A, Falletta, P, Béland, M and Seto, P (2009) Buffer

requirements for enhanced hydrogen production in acidogenic digestion of food wastes. Bioresour

Technol 100:5097-5102

10. Li, L, Langdon, AG, Nair, GR & Swan, JE (2010) Acetic acid recovery and pH control by gas stripping

and acid anaerobic sludge. Chemeca 2010, Adelaide, Australia 992-1001

11. Z Li, W Qin and Y Dai (2002) Liquid-liquid equilibria of acetic, propionic, butyric, and valeric acids

with trioctylamine as extractant. J Chem Eng Data 47:843-848

12. Li L (2013) Recovering Volatile Fatty Acids from an Acidic Anaerobic Digester, PhD Thesis,

University of Waikato, Hamilton, New Zealand


13. Nazaroff, WW and Alvarez-Cohen, L (2001) Environmental Engineering Science, John Wiley & Sons,

NY

14. Chang, JS, KS Lee, and PJ Lin (2002) Biohydrogen production with fixed-bed bioreactors. Int J

Hydrogen Energy 27:1167-1174

15. Chen, CC, CY Lin, and MC Lin (2002) Acid-base enrichment enhances anaerobic hydrogen production

process. Appl Microbiol Biotechnol 58:224-228

16. Fang, HHP, C Li, and T Zhang (2006) Acidophilic biohydrogen production from rice slurry. Int J

Hydrogen Energy 31:683-692

17. Morimoto, M, M Atsuko, AAY Atif, MA Ngan, A Fakhru'l-Razi, SE Iyuke, and AM Bakir (2004)

Biological production of hydrogen from glucose by natural anaerobic microflora. Int J Hydrogen

Energy 29: 709-713

18. Lay, JJ, KS Fan, J Chang, and CH Ku (2003) Influence of chemical nature of organic wastes on their

conversion to hydrogen by heat-shock digested sludge. Int J Hydrogen Energy 28:1361-1367

19. van Haandel, AC, and G Lettinga (1994) Anaerobic Sewage Treatment - A Practical Guide for Regions

with a Hot Climate. John Wiley & Sons, New York

20. Pantamas1, P, Chaiprasert P, and Tanticharoen M (2003) Anaerobic Digestion of glucose by Bacillus

licheniformis and Bacillus coagulans at low and high alkalinity, Asian J Energy Environ, 4:(1-2), 1-17

21. Niven, SJ, JD (2004) Beal, and PH Brooks, The simultaneous determination of short chain fatty acid,

monosaccharides and ethanol in fermented liquid pig diets. Animal Feed Sci Tech 117:339-345

22. Oude Elferink, SJWH, Krooneman, J, Gottschal, JC., Spoelstra, Faber, F and Driehuis, F (2001)

Anaerobic Conversion of Lactic Acid to Acetic Acid and 1,2-Propanediol by Lactobacillus buchneri.

Appl Environ Microbiol, 67: 125-132

23. Bilitewski, B, Härdtle, G and Marek, K (1997) Waste Management, Berlin, Springer

24. Dwidar, M, Park, JY, Mitchell, RJ and Sang, B (2012) The Future of Butyric Acid in Industry. The

Scientific World Journal

25. Campbell, NA and Reece, JB, Biology, San Francisco Pearson, Benjamin Cummings (2005).

26. McCarty, PL and Smith, DP (1986), Anaerobic wastewater treatment, Environ Sci Technol, 20: (12),

1200-1206

27. Ostrem, K, (2004), Greening Waste: Anaerobic Digestion for Treating the Organic Fraction of

Municipal Solid Wastes, MSc Thesis, Columbia University, New York.

28. Beynon, RJ and Easterby, JS (1996) Acids and Bases Buffer Solutions. Oxford Tokyo: Oxford

University Press Inc, New York. p 20

29. Siparsky, G, Voorhees, K and Miao, F (1998) Hydrolysis of polylactic Acid (PLA) and

polycaprolactone (PCL) in aqueous acetonitrile solutions: Autocatalysis. J Environ Polym Degr, 6: 31-

41

30. Zoetemeyer, RJ, Heuvel VD and Cohen A (1982) pH influence on acidogenic dissimilation of glucose

in an anaerobic digestor, Water Res, 16:303-311


Bioreactor Recovery system

Volatile fatty acids

Pump

N2

Vent

Sampling port

StirrerStirrer

Return line

Calcium carbonate slurrySludge

Fig 1. Schematic of the fed-batch/stripping/recovery system


Table 1 Products formed during the acid anaerobic digestion of glucose. The glucose added (GA) and

remaining (GR) and acid content before digestion (BD) and after digestion (AD) for repeated fed batch

digestion/stripping cycles are expressed as the mmols in the 1.7 L reactor fluid. For Cycles 2 and 3 the BD

values are those for the supernatant after stripping (AS)

Cy

cle

GA

(m

mo

l)

CH

4 (

L)

H2 (

L)

CO

2 (

L)

Sta

ge

GR

(m

mo

l)

LA

(m

mo

l )

HA

c (m

mo

l)

PA

(m

mo

l)

IBA

(m

mo

l)

BA

(m

mo

l )

IVA

(m

mo

l)

VA

(m

mo

l)

1 142 0.16 0.83 3.9 BD nd nd nd nd nd nd nd nd

AD 8.8 36.2 54.4 6.7 nd 35.4 nd nd

2 56.7 nd 0.69 2.1 AS/BD nd nd 72 6.4 1.9 22.2 nd nd

AD 3.7 5.7 71.7 4.9 2.1 46.9 0.6 nd

3 227 nd 3.0 8.9 AS/BD nd nd 84.9 3.9 1.1 13.9 nd nd

AD nd 11.9 62.8 3.3 1.0 184 0.8 4.0

4 94.4 nd nd nd

AS/BD nd nd 42.6 1.6 nd 66.6 nd 4.0

AD1* nd 251 52.9 1.8 nd 76.3 nd nd

AD2* nd 259 52.5 1.9 1.0 82.5 nd nd

nd - not detected, AD1* - sampled 19 months after glucose addition, AD2* - sampled 23 months after glucose addition, GA - Glucose added,

GR - glucose remaining, LA - lactic acid, HAc - acetic acid, PA - propionic acid, IBA - iso-butyric acid, BA - butyric acid, IVA - iso-valeric

acid, VA - valeric acid


Table 2 Comparison of observed and calculated pH values for pH before and after stripping

Cycle Stage Observed pH Calculated pH

1

AD 4.65 5.09

AS 5.12 5.01

2

AD 4.78 4.81

AS 5.01 5.01

3

AD 4.32 4.32

AS 4.95 4.92

4

AD1 3.85 3.86*

AD2 3.58 3.83*

AD1 Digestion Cycle 4 after 19 months, AD2 digestion Cycle 4 after 23 months, *Calculate using pKa of lactic acid (3.86)


Table 3 Carboxylic acid production and carbon conversion for successive digestion/stripping cycles Amounts

are expressed as either mmol (for carboxyl) or mmol C (for glucose, CO2 and acids) added to, stripped from or

present in the 1.7 L of reactor contents)

Cycle

GA

(mmol

C)

CO2

(mmol

C)a

Total carboxylb

(mmol)

Carboxyl strippedc

(mmol)

Total acid

(mmol C)

Total products

(mmol C)

AD AS

1 852 164 133 103 30 379 543

2 340 82 132 103 29 114 196

3 1362 351 268 112 156 696 1047

4 566 nd

382AD1

nm nm 809AD1

809AD1

397AD2

nm nm 861AD2

861AD2

nm – not measured, nd – not detected aCollected over water, bDetermined from the moles of all the carboxylic acids measured, cCalculated from change in COOH before and after stripping

AD – after digestion, AS – after stripping


Table 4 VFA composition of product recovered from a glucose-fed, acidic anaerobic digester

Volatile fatty acid HAc PA BA VA TVFA

Amount (mmol) 5.49 0.92 29.5 1.11 37.0

Composition (% (w/w) ) 11.4 2.3 82.9 3.5 100

HAc - acetic acid, PA - propionic acid, BA - butyric acid, VA - valeric acid, TVFA – Total volatile fatty acid

preparation of volatile fatty acid (vfa) calcium salts by anaerobic digestion of glucose

Documents