biofilm matrices as biomonitoring agent and …

Journal of Environmental Engineering & Sustainable Technology JEEST Vol. 05 No. 02, November 2018, Pages 61-67 http://jeest.ub.ac.id

P-ISSN:2356-3109 E-ISSN 2356-3117 61

BIOFILM MATRICES AS BIOMONITORING AGENT AND BIOSORBENT FOR

CR(VI) POLLUTION IN AQUATIC ECOSYSTEMS

Andi Kurniawan1, 2*

1 Coastal and Marine Research Centre, University of Brawijaya 2 Faculty of Fisheries and Marine Science, University of Brawijaya

Email: [email protected]

ABSTRACT

Heavy metal pollution in aquatic ecosystems

has become one of the primary environmental

problems. Cr(VI) is one of the most toxic and

carcinogenic heavy metal pollutants. The

degradation of aquatic ecosystems due to the

Cr(VI) pollution usually occurs slowly, but the

impact is accumulative. Hence, an awareness of

the pollution often occurs when the impacts

have already in the acute or chronic level.

Therefore, the technologies to monitor and to

solve the Cr(VI) pollution are critically

important. The application of biological

resources emerges as an alternative technology

to solve the problems. This study analyzes the

utilization of biofilm as a biomonitoring agent

and a biosorbent to monitor and to immobilize

Cr(VI) in the aquatic ecosystems, respectively.

The development of the biofilm as a

biomonitoring agent is conducted through the

investigation of Cr(VI) concentration in the

biofilm and the surrounding river water, while

the utilization of the biofilm as a biosorbent is

developed through the analysis of Cr(VI)

adsorption characteristics to the biofilm. The

results of this study reveal that the

concentrations of Cr(VI) inside the biofilms are

hundreds of times higher than the surrounding

river water. The biofilms seem to accumulate

the Cr(VI) from the surrounding river water

through a physicochemical process. According

to the result of this study, biofilms can become

a promising biological agent to monitor and to

immobilize Cr(VI) in the aquatic ecosystems.

Keywords: Biofilm; Biomonitoring;

Biosorption; Cr(VI); Water Pollution.

1. INTRODUCTION

Aquatic ecosystems have been

contaminated by various pollutants such as

heavy metal including Cr(VI) (Kyzas and

Matis, 2018; Hea, et al., 2018). The pollutants

lead to the degradation of aquatic ecosystems

which usually occurs gradually, but its effect is

accumulative. Hence, the awareness about the

degradation appears when the impacts have

been already in the chronic level (Bland, et al.,

2018; Singh, 2015).

Heavy metals such as Cr(VI) have

become a serious environmental problem

because it’s characteristics that persistent and

may be easily accumulated and magnified

through food webs (Sukandar and Kurniawan,

2017). Cr(VI) is not only toxic but also may

lead to various human health problems such as

cancer and kidney dysfunctionality (Chen, et

al., 2017). Cr(VI) largely exists in wastewater

of various industrial activities such as a leather

tanning process (Jobby, et al., 2018). The

contamination of the ion will increase along

with the increase of its utilization. Thus, the

existence of the Cr(VI) in the aquatic

ecosystems should be continuously monitored.

Various technologies to monitor the

presence of heavy metals such as Cr(VI) in the

aquatic ecosystems have been proposed

(Fomina and Gadd, 2014; Chojnacka, 2010).

One of the promising technologies is

biomonitoring. Biomonitoring is the utilization

of a living organism or a part of a living

organism as a biological agent to monitor the

pollutant in the ecosystems (García-Seoane, et

al., 2018; Kirman, et al., 2018). The primary

requirements of the biological agents are

ubiquitous, easily grown and able to accumulate

the pollutants.

The efforts to remove the excess of

Cr(VI) from the aquatic ecosystems is critically

important. Various technologies have been

proposed to solve heavy metals pollution

including Cr(VI). Biosorption is one of the

proposed technologies which have been

reported to become inexpensive and

environmentally safe. This technology uses

living organisms or a part of the living

mailto:[email protected]

Journal of Environmental Engineering & Sustainable Technology (JEEST) P-ISSN:2356-3109 Vol. 05 No. 02, November 2018, Pages 61-67

62

organisms to immobilize the pollutant from the

ecosystems. The primary key to success in the

biosorption is the ability of biosorbent to adsorb

the pollutant. The biological agents used as a

biosorbent should be easy to be obtained and

having a high capacity to remove the pollutant.

Biofilm as a predominant habitat of

microbes (Flemming and Wingender, 2010;

Costerton, et al., 1995; Fabbri, et al., 2018) can

become a biomonitoring agent and biosorbent

for Cr(VI) in the aquatic ecosystem. Biofilm is

a matrix-enclosed microbial community that

attached on the almost every substrate in the

aquatic ecosystems (Tsuchiya, et al., 2009).

Previous studies have shown that the biofilm

can attract various ions including heavy metals

(Kurniawan, 2012; Kurniawan, 2015). In this

study, the Cr(VI) concentration in the biofilm

and the surrounding water of the biofilm in the

river ecosystems were investigated in order to

analyze the biofilm utilization as a

biomonitoring agent. Moreover, the

characteristics of Cr(VI) adsorption to the

biofilm were also analyzed to develop the

biofilm as a biosorbent for Cr(VI). The results

of this study suggest that the biofilm can

become promising biomonitoring agent and

biosorbent to monitor and remove the heavy

metal such as Cr(VI) from the aquatic

ecosystems.

2. MATERIALS AND METHODS

2.1. Sampling Site And Sample

Preparation

Samples in this study were biofilms

formed on the stones and the surrounding river

water of the biofilms (ca. 15 cm from the

biofilm surface) collected from the Badek River

in Malang City, East Java, Indonesia. In the

around riverbank area of the Badek River, there

are leather tanning plants. The plants often

discard the wastewater from the leather tanning

process to the Badek River. The samples of the

biofilms were collected from 3 sampling sites.

Site 1 is the location near the outlet of the

tannery plants, Site 2 is in the middle of the

watershed area between the Site 1 and the end

of the watershed of Badek River, and Site 3 is

at the end of the Badek River watershed before

the river joined with the Brantas River.

The samples of biofilms and the

surrounding water were collected in 2 times.

The first sampling time is when the tannery

plants operated, and the second sampling times

is when the tannery plants did not operate. The

samples were collected from the middle and the

both sides of the river.

The stones covered by biofilm and the

surrounding river waters were brought back to

the laboratory in plastic containers. The

temperature of the container is maintained at

around 4 °C. The biofilms were removed from

the stones using a sterilized toothbrush in 80 mL

of distilled water. The Cr(VI) concentration in

the suspension was assumed as the diluted

Cr(VI) concentration inside the biofilms. The

Cr(VI) concentrations were measured using an

Atomic Absorption Spectroscopy Shimadzu

AA-6800 (Shimadzu Corporation, Japan). All

the samples were stored in - 25 °C before used

in the experiment.

2.2. Kinetics Of Adsorption

Eighty milliliters of 50 mg·L⁻¹ K₂Cr₂O7

aqueous solutions were prepared by diluted

reagent grade K₂Cr₂O7 to the distilled water. 0.5

gram of biofilm was added to the suspension

and mixed well using a magnetic stirrer. The

biofilm suspensions were sub-sampled after 5,

15, 30, 60, and 180 minutes. The sub-sample

was centrifuged (8,000 × g at 4°C for 10 min)

and the supernatant was collected. The

concentration of Cr(VI) in the supernatants

were measured using an Atomic Absorption

Spectroscopy Shimadzu AA-6800 (Shimadzu

Corporation, Japan). The same K₂Cr₂O7

aqueous solutions without the biofilm were

used as a control in the kinetics of adsorption

experiment. The adsorbed amounts of Cr(VI)

were calculated from the difference of the

Cr(VI) concentration in the control and the sub-

samples. All the kinetics experiments were

repeated three times independently.

2.3. Adsorption Isotherm

A half gram of biofilm pellet was added

to 80 mL of K₂Cr₂O7 aqueous solution (5, 25,

50, 300, 600, 1200 mg·L⁻¹) prepared with the

same method described above. After 15

minutes, the suspensions were centrifuged

(8,000 × g at 4°C for 10 min), and then, the

supernatants were collected. As the control, the

same concentration of K₂Cr₂O7 aqueous

Andi Kurniawan, Biofilm Matrices As Biomonitoring Agent And Biosorbent …

P-ISSN:2356-3109 E-ISSN 2356-3117 63

solutions without the addition of the biofilm

was prepared. The adsorbed amounts of Cr(VI)

to the biofilms were calculated from the

difference of concentration between the control

and the supernatants. The concentration of

Cr(VI) was measured using an Atomic

Absorption Spectroscopy Shimadzu AA-6800

(Shimadzu Corporation, Japan). All the

adsorption isotherm experiments were repeated

three times independently.

The adsorption isotherm of biofilm is

analyzed using the variant of the Langmuir

Adsorption Equation as described below

(Vijayaraghavan and Balasubramanian, 2018;

Freifelder, 1985)

maxmax )(

1

N

C

bNN

C　

(1)

The Langmuir adsorption equation assumes that

a dynamic equilibrium exists between the

adsorbed Cr(VI) (N; mg·g⁻¹) and the free

Cr(VI) in solution (C; mg·L⁻¹). The adsorption

equilibrium constant (b) is the ratio of the

adsorption and desorption rates. The plot of C/N

against C shows a straight line with a slope of

1/Nmax and a y-axis intercept of 1/(Nmax)b.

Then, the values of Nmax (the maximum

amount of adsorbed ion; mg·g⁻¹) and b (L·mg⁻¹)

were calculated (Kurniawan, 2012).

3. RESULTS AND DISCUSSION

The concentration of Cr(VI) inside the

biofilms and the surrounding water were

investigated in this study. The characteristics of

Cr(VI) adsorption to the biofilms were also

analyzed. The results are used to examine the

possibility to utilize the biofilm matrices as a

biomonitoring agent and a biosorbent for

Cr(VI) contamination in the aquatic

ecosystems.

3.1. Cr(VI) Concentration In The

Biofilm And The Surrounding

Water

The concentrations of Cr(VI) in the

biofilms and the surrounding river waters were

measured when the leather tanning plants did

not operate and operated (Table 1). The results

indicate that the concentration of Cr(VI) inside

the biofilms were hundred times higher

compared to those of in the surrounding river

waters. This results indicated that biofilms

might accumulate the Cr(VI) from the

surrounding waters [16]. Biofilms in the aquatic

ecosystems have been reported to have the

ability to attract and reserve ions including

heavy metal ions such as Cr(VI) (Kurniawan,

2015; Cheng, et al., 2018, Julien, et al., 2014).

When the tannery plants operated,

concentrations of Cr(VI) in the water of Badek

River were 0.078-0.13 mg·L⁻¹. These

concentrations are higher than the threshold for

human health (0.05 mg·L⁻¹). These facts

suggested that the Badek River seems to be

contaminated by the Cr(VI). However, the

concentrations of Cr(VI) in the river water when

the tannery plants did not operate were only

0.022-0.041 mg·L⁻¹ which are lower than the

threshold of human health. These results mean

that if the measurement to investigate the

Cr(VI) contamination in the river conducted in

the time when the tannery plants did not

operate, the contamination of Cr(VI) in the

Badek River will not be revealed. These

conditions may lead to the misjudgment on the

river pollution status. This fact revealed the

limitation of the utilization of Cr(VI)

concentration in the river water as a standard to

monitor the water pollution.


64

Table 1. The concentration of Cr(VI) inside the biofilm and in the surrounding river waters.

Experiment repeated 3 times. ± values represent the standard deviation.

The monitoring of heavy metals such as

Cr(VI) contamination in the river requires the

more suitable and accurate indicators than the

concentration of the heavy metals in the river

water. Hence, the concentration of Cr(VI) in the

indicators should represent and indicate the

existence of the heavy metals in the river not

only in the sampling day but also in the

extended periods. Heavy metals such as Cr(VI)

are also can be accumulated in the living

organism, and then, through the food web can

be magnified. Therefore, the indicators should

also represent the potentiality of this dangerous

cycles. In this case, the utilization of biological

agents to monitor the heavy metal pollution

may appear as a promising alternative.

Biofilm is the predominant habitat of

microbes and ubiquitous in the aquatic

ecosystems. The biofilm easily forms on the all

substrates in the aquatic ecosystems [13].

Biofilm also directly involves with the food

chain because it is a feed for many fishes, and

thus, the heavy metals accumulated inside the

biofilm may be magnified through the food

webs. The results of this study show that the

biofilms may accumulate the heavy metals such

as Cr(VI) from the surrounding waters. If there

is a heavy metal input into the river at a

particular time, the heavy metal will be attracted

and retained in the biofilm matrix for an

extended period. Hence, the presence of heavy

metals in the biofilm is very likely to be used as

an indicator of heavy metal input into the river.

Moreover, the heavy metal

concentrations inside the biofilm will

dynamically change along with the change of

the ion concentrations in the surrounding waters

[17]. When the concentration of the heavy

metals in the surrounding water decrease, the

biofilm may release the heavy metal to the

surrounding water. The results of this study

suggest that the biofilm may be used as a

promising biological agent to monitor heavy

metal pollution in the aquatic ecosystems.

3.2. Kinetics Of Adsorption

The time course of Cr(VI) adsorption to

the biofilm was investigated (Fig. 1). The

adsorbed amounts of the Cr(VI) to the biofilm

were relatively stable from 5 min until the end

of the experiment (± 1,8 mg·g⁻¹). This result

suggested that the adsorption of Cr(VI) to the

biofilm is a speedy process. This process is a

characteristic of the passive uptake mechanism

(Kyzas, et al., 2014). It seems that the

accumulation of Cr(VI) to the biofilm in this

study occurred through a physiochemical

process (Kurniawan, 2015; Ifelebuegu, et al.,

2018). Our previous studies suggested that the

biofilms may attract and retain ions including

heavy metals such as Cr(VI) through the ion

exchange mechanism and the electrostatic

interactions (Kurniawan, 2012).

Not operating 3.2 ± 0.23 2.04 ± 0.07 1.5 ± 0.26

Operating 4.02 ± 0.33 3.06 ± 0.11 2.09 ± 0.14

Not operating 0.041 ± 0.01 0.033 ± 0.01 0.022 ± 0.01

Operating 0.13 ± 0.01 0.092 ± 0.01 0.078 ± 0.01

Site 2 Site 3

Cr(VI) concentration (mg·L⁻¹)

Biofilm

River Water

Sample TimeSite 1


P-ISSN:2356-3109 E-ISSN 2356-3117 65

Figure 1. Time course of Cr(VI) adsorption to

the biofilm. Experiment repeated 3

times. ± values represent the standard

deviation

3.3. Adsorption Isotherm

In order to analyze the Cr(VI) adsorption

mechanism to the biofilm in more detail, the

adsorption isotherm of Cr(VI) to the biofilm

collected from the Badek River was

investigated. The initial concentrations used in

the experiment were 5-1200 50 mg·L⁻¹. The

adsorbed amounts of Cr(VI) to the biofilm

increase along with the increase of the Cr(VI)

concentration in the surrounding waters, and

then, leveled off in the high concentration (Fig.

2).

Figure 2. Time Adsorption isotherm of Cr(VI)

to the biofilm. Experiment repeated 3

times. ± values represent the standard

deviation

The adsorption isotherm of Cr(VI) to the

biofilm shows the L type of adsorption. This

type of curve suggested the adsorption may be

described using the Langmuir Adsorption

Equation. The variant of the Langmuir

adsorption equation was used to analyze the

adsorption of Cr(VI) to the biofilms. The result

showed that the adsorption of Cr(VI) to the

biofilm fit well with the Langmuir adsorption

model (R² = 0.95) (Fig. 3). The Nmax of the

Cr(VI) to the biofilm collected from the Badek

River in this experiment is 9.09 mg·g⁻¹. The

adsorption equilibrium constant is 0.01 mg·L⁻¹.

The adsorption of Cr(VI) to the biofilm seems

to occur in a monolayer form where the retained

mechanisms are solely due to the interaction

between the Cr(VI) and the biofilm. In this case,

the binding sites in the biofilm have an equal

affinity to the Cr(VI). The adsorption of Cr(VI)

to the biofilm increase linearly in the low

concentration, then slightly increase along with

the increase of the concentration and finally

reached the saturation point where the

adsorption amount becomes constant. The

results support the suggestion that Cr(VI) seems

to be adsorbed to the biofilm through the

physicochemical process such as an ion

exchange mechanisms and an electrostatic

interaction [(Kurniawan, 2012).

Figure 3. Time The overlay of the Cr(VI)

adsorption data to the biofilm using

the variant of the Langmuir

Isotherm equation. C is the

equilibrium concentration (mg·L⁻¹)

and N is the adsorbed amount

(mg·g⁻¹)

The effectiveness of the biofilm as the

biosorbent for the Cr(VI) was investigated by

calculating the percentage of the adsorbed

Cr(VI) in the various initial concentrations (Fig.

4). The results suggested that the high

effectiveness of the adsorption occurs in the low

concentration, and then, decrease along with the

increase of the Cr(VI) concentrations in the

0

1

2

3

0 50 100 150 200

Ad

sorb

en

t a

mo

un

t (m

g·g

⁻¹)

Contact time (minute)

0

2

4

6

8

10

12

0 200 400 600 800 1000 1200

Ad

sorb

en

t a

mo

un

t (m

g ·

g⁻¹

)

Equilibrium Concentration (mg · L⁻')

y = 0.11x + 8.26

R² = 0.95

0

20

40

60

80

100

120

140

0 500 1000 1500

C/N

C (mg·L⁻¹)


66

surrounding waters. The high effectiveness in

the low concentration it seems due to the high

ratio of the available binding sites in the

biofilms compared to the amount of ions in the

surrounding waters. The effective level of

adsorption ability (the effectiveness is more

than 50 %) is reached until 50 mg·L⁻¹ of Cr(VI).

This concentration should be taken as one of the

primary considerations to develop a biofilm

reactor to remove Cr(VI) from the aquatic

ecosystems.

Figure 4. Time The effectiveness of Cr(VI)

adsorption to the biofilm

4. CONCLUSIONS

This study analyzes the utilization of

biofilm as a biomonitoring agent and a

biosorbent to monitor and immobilize Cr(VI) in

the river ecosystems, respectively. The results

show that the Cr(VI) concentration inside the

biofilm is hundred times higher compared to the

Cr(VI) concentration in the river water. The

Cr(VI) concentration inside the biofilms can

represent the input of the ions to the river

ecosystems for an extended period, and thus,

represent the history of Cr(VI) input to the

ecosystems. This study revealed that the biofilm

matrix might become a promising biological

agent to monitor water pollutant such as Cr(VI)

in the aquatic ecosystems. The biofilms of River

Badek seems to adsorb Cr(VI) through

physicochemical interactions. The maximum

Cr(VI) adsorption ability of the biofilm is 9.09

mg·g⁻¹ and the adsorption equilibrium constant

is 0.01 mg·L⁻¹. According to the results of this

study, the biofilm may become a promising

biomonitoring agent and a biosorbent for water

pollutant such as Cr(VI) in the aquatic

ecosystems.

FUNDING

This research is supported by the Directorate for

Research and Community Service, Directorate

General of Strengthening Research and

Development, Ministry of Research,

Technology and Higher Education of the

Republic of Indonesia.

CONFLICTS OF INTEREST

The author declares no conflict of interest

related to this study.

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