© 2021 The Authors Water Science & Technology Vol 84 No 7, 1804 doi: 10.2166/wst.2021.

377

Determination of kinetic coefficients for treating synthetic oily wastewater

in suspended growth batch fed reactor

P. Sanghamitraa,*, Debabrata Mazumder a and Somnath Mukherjeeb

a
Civil Engineering Department, Indian Institute of Engineering Science and Technology, Shibpur, Howrah-711103, West Bengal, India
b
Civil Engineering Department, Jadavpur University, Kolkata 700032, West Bengal, India
*Corresponding author. E-mail: sanghamitra.poly@gmail.com

ABSTRACT

Discharge of oily wastewater imparts serious threat to the environment because of high level concentration of chemical oxygen demand (COD),
biochemical oxygen demand (BOD) as well as oil and grease and it is difficult to treat such wastewater due to its inherent toxic and inhibitory
property. A treatability study of oily wastewater (carrying petroleum) has been performed in the present work using a batch suspended growth
reactor. The experiment was conducted using acclimatized suspended biomass in laboratory environment and the kinetic coefficients were
determined which are immensely important for the design of such reactor. The oil removal efficiency was observed to be in the range of
62.84–85.45% corresponding to average MLSS concentration range of 1,797–3,668 mg/L. Haldane kinetic model was found to be the best
fitted for the biodegradation of oily wastewater with acclimatised microorganisms in the present investigation. The kinetic co-efficients includ-
ing Ks, Y, kd, k and ki were calculated from the experimental data and the values were compared with published results cited by various
scientists. The derived kinetic coefficients values are to be useful for understanding the dynamics of substrate utilisation with production of
biomass and efficient design of biological systems and also for pilot plant investigation with real life wastewater of similar nature.

Key words: batch suspended growth reactor, Haldane equation, kinetic coefficients, oil and grease removal

HIGHLIGHTS

• Biological treatment of petroleum based synthetic oily wastewater which poses inhibitory effect.
• Determination of kinetic coefficients for aerobic treatment of such wastewater in suspended growth batch fed reactor along with perform-
ance study using Haldane’s approach.
• In the present work, oil and grease was selected as performance parameter instead of COD (which is commonly used by most of the
researchers).

INTRODUCTION
The blooming economy, massive urbanization and mushrooming growth of industries emerge substantial amount of oily
wastewater in the water environment. Such typical oily wastewater emanates from petrochemical industries, oil refineries,
garages or workshops, kitchens, restaurants and other industries meant for processing food, beverage and vegetable oil,
etc. (Yu et al. 2017; Sanghamitra et al. 2021). Discharge of such oil containing wastewater leads to numerous environmental
problems, viz. reduced crop growth, pollution of surface and ground water, soil pollution, sewer line blockage and hence,
needs appropriate treatment for the purpose of complying environmental regulations and in anticipation of reuse and recy-
cling of such wastewater. Though the discharge of oily wastewater contributes inhibitory environment, such problem can be
abated by using well acclimatised biomass which are able to degrade oil and grease from such wastewater. Since rate of degra-
dation is slow under the environment of oil and grease, bacteria should be grown carefully and the retention time necessary
for the growth should be observed.
For appropriate design of biological reactors as well as optimization of operational process parameters, it is very much
essential to acquire the knowledge of biokinetics (Nakhla & Al-Harazin 1993; Contreras et al. 2001; Nakhla et al. 2006).
The factors which are indispensable in governing kinetics include the rate of utilization of substrate and microbial growth
rate, i.e yield rate which have been utilized by many researchers in the earlier studies (Sanghamitra et al. 2019). These
two above factors have been considered in various models, i.e. Monod, Haldane and Aiba-Edwards for determination of

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 Water Science & Technology Vol 84 No 7, 1805

kinetic parameters. In both Monod and Haldane kinetic model, along with the increase in substrate concentration, the
growth rate increases and after maximum growth rate, asymptotic approach is observed in Monod’s model while there is
decline in growth rate after peak in Haldane’s model (Rozich & Colvin 1986; Kim et al. 2005). Though Monod kinetic
model has been widely used in the literature, Haldane kinetic model was also extensively used in case of inhibitory waste-
water with high strength (Nakhla et al. 2006).
Most of the researchers have done performance and kinetic evaluation of the reactors treating edible oily wastewater con-
sidering mainly chemical oxygen demand (COD) and sometimes components of oil, i.e. phenol as performance parameter.
Ahmadi et al. (2006) studied kinetic behaviour for the treatment of olive mill wastewater for the removal of phenolic pollu-
tants which followed Monod kinetics. Again, Nakhla et al. (2006) applied kinetic model for aerobic biodegradation of pet
food wastewater in activated sludge reactor (batch type) for removal of COD. The data of substrate as well as biomass
were fitted better in Haldane model than Monod model. Furthermore, Kul & Nuhoğlu (2020) observed biodegradation of
olive mill wastewater for COD removal by mixed cultures in an aerobic batch reactor and studied inhibition models for evalu-
ation of kinetic parameters. Haldane model was observed to be the best tested model in this work.
Very few works are published for the biodegradation and estimation of kinetic parameters of petroleum carrying oily waste-
water in terms of COD. Rahman & Al-Malack (2012) studied the kinetics of a cross flow membrane bioreactor (lab scale) for
treating refinery wastewater at mixed liquor suspended solids (MLSS) concentration of 3,000 and 5,000 mg/L for COD removal
by using Monod model. The kinetic coefficients obtained in this study were found to be in the normal range except the yield co-
efficient (Y). The kinetics was also studied for the degradation of crude oil by microbial species Pseudomonas aeruginosa by
using Monod kinetics for rate prediction of biological treatment of oily wastewater (Talaiekhozani et al. 2015).
In order to design and understand the biological treatment process, kinetics is immensely useful (Kaewsuk et al. 2010). The
importance of using various biokinetic parameters are explained in various literatures. The change in specific growth rate of
bacteria along with the variation in concentration of growth limiting substrate can be accomplished by half saturation con-
stant (Ks) value. The volume of various biological reactors can be obtained by evaluating maximum specific substrate
utilisation rate (k) value as the size of the reactor increases with decrease in value of k. The total quantity of sludge produced
during wastewater treatment can be estimated from yield coefficient (Y). Furthermore, the total quantity of sludge needed to
be handled, the cost and size of sludge handling chamber can also be determined from endogenous decay constant (kd) value
(Prakash 2013).
Several kinetic models have been studied earlier for suspended growth reactor system which is described in literature for
treatment of urban sewage and various biodegradable industrial effluents. But no such information till date is much available
for biological treatment of oily wastewater including its biodegradation kinetics specifically for evaluation of various kinetic
coefficients for non-edible petroleum based oily wastewater, which is slowly biodegradable and poses inhibitory effect during
treatment process. The authors felt estimation of bio-kinetic parameters is essentially needed for aerobic treatment of such
wastewater for the rational design of aeration tank. Furthermore, most of the researchers considered COD as performance
parameter instead of taking initial oil and grease concentration as governing input functions during kinetic study of oily
wastewater. The objective of the present investigation was to study the performance of a laboratory scale batch suspended
growth aerobic reactor for treatment of petroleum based synthetic oily wastewater to derive kinetic coefficients using the
experimental data for substrate utilization as well as biomass production with the help of Haldane kinetic model.

MATERIALS AND METHODS

Acclimatization of biomass
A laboratory scale batch fed reactor (bucket type) of capacity 5 L and working volume of 3 L was used in this study. The reac-
tor content was aerated using two aqua pumps. The reactor was running in Environmental Engineering Laboratory, Civil
Engineering Department, IIEST, Shibpur for removal of oil and grease from synthetic oily wastewater. The microorganisms
acclimatized in the present work was isolated from the soil samples collected from petroleum waste disposal sites near the
above mentioned Institute using synthetic dextrose feed (Feed 1) with concentration 10,000 mg/L as well as oil feed (Feed 2)
with concentration 1,000 mg/L. The C:N:P ratio for both feed solutions are 100:5:1. Acclimation process was continued in
the reactor for a time period of three months unless a steady condition was arrived (Sanghamitra et al. 2020). Tables 1 and 2
exhibit the composition of edextrose and oil feed respectively. pH of the reactor was maintained in the range of 7.2–8 during
acclimatization.

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 Water Science & Technology Vol 84 No 7, 1806

Table 1 | Composition of synthetic dextrose bearing wastewater (Feed 1)

Constituents Amount added per litre in dextrose bearing feed

Dextrose 10 g
NH4Cl 0.7642 g
KH2PO4 0.1754 g

Table 2 | Composition of synthetic oily wastewater (Feed 2)

Constituents Amount added per litre in oil bearing feed

Oil (Castrol CRB prima diesel engine oil) 1.11 mL

NH4Cl 0.1364 g
KH2PO4 0.0313 g

Experimental procedure for batch kinetic study

A sizeable number of batch studies were conducted after successful acclimatization of biomass by using synthetic oily waste-
water. During batch study, sample of 50 mL was collected from the batch suspended growth reactor at a certain time interval.
The initial concentration of oil in samples for different sets during batch performance in non-inhibition zone is shown in
Table 3. Various parameters, i.e. pH, mixed liquor suspended solids (MLSS) and oil and grease, have been measured by
using the procedure as laid down in Standard Methods (APHA 2000). Measurement of pH was done by using pH meter
(Orion make) before sample filtration, MLSS by filtering and drying the biomass present in sample in hot air oven at 105 °
C using commercial filter paper and oil and grease by Soxhlet Extraction method.

Kinetic model study

During batch experiments, the decrease in concentration of substrate was observed, by noting the decrease in substrate util-
ization rate beyond a certain substrate concentration. This trend clearly indicated there was a substrate inhibition, which was
required to be expressed in form of a suitable ‘inhibition model’. Accordingly, all the relevant data need to be fitted in a par-
ticular model in order to obtain bio-kinetic coefficients (Simkins & Alexander 1984; Nakhla et al. 2006). Haldane kinetics
would be a viable and realistic option under the present condition, which had been widely used for inhibition dynamics
on account of compatibility and simplicity (Keesman & Stigter 2003). In addition, substrate inhibition characteristics were
taken into consideration in this model.
As per Nakhla et al. (2006), the average percentage error for Monod model as well as Haldane model were found to be
12–154% and 3–33.3% respectively for biomass and substrate concentrations. It was further revealed by applying paired

Table 3 | Initial concentration of oil in the synthetic samples during batch study

Batch no Initial concentration of oil (mg/L)

1 69
2 78
3 80
4 81
5 91
6 92
7 99
8 104
9 110

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 Water Science & Technology Vol 84 No 7, 1807

t-test in case of Haldane model was superior to Monod model. Again, for Monod’s model, the average values of kinetic coef-
ficients varied more widely than Haldane’s model, hence more consistent and stable kinetic coefficients were obtained by the
application of Haldane’s model. Therefore, Haldane’s kinetic approach was applied in the present work for determination of
kinetic coefficients. It was also reported that Monod’s model was not enough for modelling the organic compounds with inhi-
bition potential and the best result was obtained by using Haldane model during treatment of olive mill wastewater in aerobic
batch reactor (Kul & Nuhoğlu 2020).

Evaluation of kinetic coefficients

The relation between specific growth rate with substrate concentration as per Monod’s model is represented by Equation (1).

mm S
m¼ (1)
Ks þ S

Specific growth rate as per Haldane’s model is represented by Equation (2).

mm S
m¼ (2)
Ks þ S þ (S2 =Ki )

where, μm is maximum specific growth rate (d1)

S is concentration of oil and grease at any time (mg/L)
Ki is inhibition constant (mg/L)
In case of Haldane’s kinetics, the specific growth rate increases with the increase in substrate concentration up to peak value
of growth rate (non-inhibition zone), after which growth rate declines with respect to the increase in substrate concentration
(inhibition zone). The batch study data within non-inhibition zone was considered for determination of kinetic parameters.
The value of k and Ks can be determined from substrate utilization kinetics as represented in Equation (3).

QXavg Ks 1 1
¼ : þ (3)
S0  S k S k

The value of Y and kd can be determined from biomass growth kinetics as represented in Equation (4).

X0  X S0  S
¼ :Y  kd (4)
QX QXavg

Here, S0 is initial concentration of oil in the reactor (mg/L)

S is final concentration of oil in the reactor (mg/L)
X is concentration of biomass (mg/L)
Ө is batch period (d)
X’ is final concentration of biomass over time Ө (mg/L)
Xavg is average biomass concentration (mg/L).

RESULTS AND DISCUSSION

Batch study was carried out using synthetic oily wastewater with the concentration of oil in the range of 69–161 mg/L. During
batch studies, pH, MLSS and oil and grease were measured at time interval of 24 to 120 h depending on batch performance.
The results of batch study for pH and oil and grease used for determination of kinetic constants are shown in Figures 1 and 2,
respectively.
The initial value of pH in different batches varied in the range of 7.6–8.07 during the experimental period of around four
months by addition of synthetic oily wastewater of concentration 1,000 mg/L with C:N:P ratio of 100:5:1 as shown in Figure 1.
As pH was found changing during the process of biodegradation proceeds, the range of pH varies from one batch to another
accordingly. In batch 2, 4 and 8, no significant change of pH was observed. During batch study, pH was maintained in the

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 Water Science & Technology Vol 84 No 7, 1808

Figure 1 | Profile of variation of pH during batch study.

Figure 2 | Profile of variation of residual oil during batch study.

range of 7.2–8 by addition of 1:10 HCl and 1N Na2CO3. pH adjustment is essential for survival and appropriate growth of
biomass in order to achieve better treatment efficiency (Fulazzaky et al. 2020). Figure 2 represents the trend of oil in different
batches in which the concentration of oil decreases with the progress of the batch period.
The percentage removal of oil was in the range of 62.84–85.45% as shown in Figure 3. Such removal efficiency of oil during
batch study was obtained in correspondence to average MLSS concentration. within the range of 1,797–3,668 mg/L. The
maximum oil removal of 85.45% was obtained with respect to an initial oil concentration of 110 mg/L for hydraulic retention
time (HRT) of 220 h and corresponding average MLSS concentration of 3,668 mg/L. At an initial oil concentration of 69 mg/
L, minimum oil removal of 62.84% was achieved against an average MLSS concentration of 1,797 mg/L. The results reveal
that oil removal is both initial concentration of oil and microbial population dependent. It was also observed from the batch
study results that oil removal was significantly high during 96–270 h in different batches.
The specific growth rate of biomass was calculated for different batches and was plotted against substrate concentration as
shown in Figure 4. This graph shows increase in specific growth rate with increase in the concentration of substrate and it
decreases after reaching peak point which implies Haldane kinetics is applicable in such system.
Various kinetic parameters, i.e. Ks, k, Y, kd and ki, were obtained under Haldane’s model. The experimental data during
batch study was considered for calculation of Ks and k in non-inhibition zone, i.e. within low concentration of oil and

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 Water Science & Technology Vol 84 No 7, 1809

Figure 3 | Percentage oil removal efficiency during batch study.

Figure 4 | Specific growth rate versus substrate concentration.

grease. The values of [(ӨXavg)/(S0-S)] were plotted against 1/S for determination of Ks and k as represented in Figure 5. Again,
[(X0 -X)/(ӨX)] was plotted against [(S0-S)/(ӨXavg)] for the determination of Y and kd as shown in Figure 6. It was observed from
the slope and intercept of Figures 5 and 6 that k ¼ 0.0197/d, Ks ¼ 96.5 mg/L, Y ¼ 0.75 mg/mg, kd ¼ 0.0012/d. Here Ki corre-
sponds to 120–161 mg/L.
The values of Ks, k, Y, kd as obtained from the linearized plots (Figures 5 and 6) and ki along with published values reported
by several researchers with kinetic model are exhibited in Table 4.

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 Water Science & Technology Vol 84 No 7, 1810

Figure 5 | Linear plots for determination of k and Ks.

Figure 6 | Line-weaver Burk plot for determination of Y and kd.

The Ks value in the present work indicates that biomass could grow well at moderate concentration which is less significant
to the values reported by Nakhla et al. (2006) and Ahmadi et al. (2006), in their earlier works but more than the value given by
Kul & Nuhoğlu (2020). This may be due to the reason that the biomass characteristics and the surrounding environment
govern half velocity constant (Walker 1998). Talaiekhozani et al. (2015) reported that rate of substrate utilization is high
at a low concentration in the case of low value of Ks and microorganisms of high concentration is necessary for increasing
the substrate utilization rate in the case of high value of Ks. Again, the value of k obtained in this work is less than the values
reported by other researchers. Furthermore, the yield coefficient (Y) in the present work was found to be 0.75 which is less
than the values mentioned by Talaiekhozani et al. (2015) obtained during treatment of crude oil, but the same as the value
obtained by Ahmadi et al. (2006) for the treatment of olive mill wastewater (OMW) for percentage concentration of OMW as
30. But the value of Y was observed to be higher than the values mentioned by various other researchers as represented in
Table 4. The value of ki indicates that inhibitory effect was started at a lesser concentration. Again, the value of kd is not cor-
roborating with other workers. The incongruity in the value may be due to dissimilarity in substrate concentrations, enzyme
limitation and experimental constraints, at the same time with differences in the principal microbial species along with the
degree of adaptation of the microbial cultures to the oily wastewater (Kul & Nuhoğlu 2020).

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 Water Science & Technology Vol 84 No 7, 1811

Table 4 | Comparison of various kinetic parameters during treatment of oily wastewater

Y (mg VSS/mg
1 1
Type of oily wastewater (model used) k (d ) KS (mg/L) substrate) kd (d ) Ki (mg/L) References

Pet food processing wastewater (Monod kinetics) 0.168 – – – – Liu et al. (2004)
Pet food wastewater (DAF pretreated-Haldane 1.28–5.35 17,833–28,477 0.13–0.51 – 48,175 Nakhla et al. (2006)
kinetics)
Pet food wastewater (raw wastewater-modified 1–1.3 5,580–5,600 0.08–0.85 – – Nakhla et al. (2006)
hydrolysis Monod model)
Olive oil mill wastewater (COD removal for % – 6,900,11,000, 2.3, 0.75, – – Ahmadi et al.
conc. Of OMW 20, 30 and 50) 8,800 0.31 (2006)
(Monod equation)
Refinery wastewater (for MLSS 2.365 396.62 0.276 0.07 Rahman & Al-
5,000 mg/L-Monod model) Malack (2012)
Refinery wastewater (for MLSS 5.4 659.45 0.222 0.09 Rahman & Al-
3,000 mg/L-Monod model) Malack (2012)
Crude oil (Monod model) 9.31 162.4 0.8896 0.1284 – Talaiekhozani et al.
(2015)
Olive mill wastewater (Haldane kinetics) 45.34 207.97 Kul & Nuhoğlu
(2020)
Synthetic oily wastewater-petroleum based 0.0197 96.5 0.75 0.0012 120–161 Present work
(Haldane model)

CONCLUSIONS
Biodegradation of oil and grease carrying synthetic wastewater was performed in a laboratory scale batch suspended growth
reactor with increasing concentration of synthetic oil feed and acclimatised biomass for the development of process kinetics.
The treatment efficiency was observed to be maximum (i.e. 85.45%) in terms of removal of oil and grease at an initial oil con-
centration of 110 mg/L with 220 h of retention time and average MLSS concentration of 3,668 mg/L. The inhibition takes
place as indicated in descending growth rate after a certain concentration of oil and grease and complying with Haldane
growth kinetics. The biodegradation of synthetic oily wastewater (petroleum based) is found to be slow and mild inhibitory
effect as exhibited the values of kinetic coefficients. Kinetic constants were obtained based on Haldane’s approach using
experimental data. The values of bio-kinetic parameters are estimated to be k ¼ 0.0197/d, Y ¼ 0.75 mg/mg, Ks ¼ 96.5 mg/L,
kd ¼ 0.0012 d1 and ki ¼ 120–161 mg/L. This research work will be useful in understanding the kinetic behaviour and biode-
gradability of batch suspended growth reactor in treating petroleum carrying synthetic oily wastewater. This will also help in
design of bioreactors for the treatment of wastewater containing oil and grease with similar characteristics.

DATA AVAILABILITY STATEMENT

All relevant data are included in the paper or its Supplementary Information.

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First received 30 June 2021; accepted in revised form 28 August 2021. Available online 14 September 2021

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