Journal of Engineering Science and Technology

Vol. XX, No. Y (Year) PPP - QQQ

© School of Engineering, Taylor’s University

TREATMENT OF HIGH-PHOSPHATE (K2HPO4) WASTEWATER

IN NORTH COLOMBIA USING CHLORELLA VULGARIS
MICROALGAE

JORGE SÁNCHEZ-APONTE1, ILDEFONSO BALDIRIS-NAVARRO1,

ÁNGEL D. GONZÁLEZ-DELGADO2,*

1
International Nautical River and Port Centre - SENA, Cartagena, Colombia.
2
Chemical Engineering Department, Nanomaterials and Computer Aided Process
Engineering Research Group (NIPAC), University of Cartagena, Cartagena,
Bolivar, Colombia
*Corresponding Author: agonzalezd1@unicartagena.edu.co

Abstract
This study analysed the capacity of the microalgae Chlorella Vulgaris to
eliminate phosphates, as well as the effect it has on the growth of microalgae. It
was found that high concentrations of K2HPO4 (10,000 mg/L and 40,000
mg/L) inhibited microalgae growth, and it was shown that there is no statistical
difference between phosphate removal at high and low concentration. Chlorella
Vulgaris is a microorganism that can remove phosphates at different
concentrations in wastewater.
Keywords: Phosphate removal, Chlorella Vulgaris, microalgae, wastewater.

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2 Sánchez-Aponte, J. et al.

1. Introduction
Microalgae have great ecological importance due to their biodiversity, high
growth rate (double their size in 24 hours), easy to grow, efficient carbon dioxide
fixation and does not required arable land to growth [1]. They are present in any
aquatic environment where there is a source of carbon, nutrients, sufficient light
and appropriate temperatures according to the species. They provide the
biosphere with a considerable amount of oxygen by fixing more than 40% of the
earth’s coal. Microalgae are very attractive to produce various products of
commercial interest since they can use residual effluents as a source of nutrients.
The use of microalgae in wastewater treatment has been the subject of numerous
researches due to its ability to remove significant amounts of nitrogen (N),
phosphorus (P), heavy metals, fats and at the same time accelerating inactivation
of pathogenic bacteria [2–4]. Microalgae-based sorbents have received much
attention because of their abundant availability in the natural environment and
their adaptability to a variety of habitats, for example, fresh and marine water,
domestic and industrial effluents and man-made wetlands [5].
The bioremediation capacity of microalgae in wastewater has been
demonstrated previously, relevant aspects such as the conversion of nutrients
present in algal biomass are already described [6–9]. Microalgal biomass can be
used in the production of bioenergetic alternatives such as biodiesel, biohydrogen
or methane [2], it can also be used as a source of pigments and nutritional
supplements [10,11], and as a adsorbent of dyes [12]. For this reason, the
production of microalgae using wastewater as a culture medium presents an
alternative for the environmental problem of the deterioration of our bodies of
water. At the same time, different objectives are achievable within the same
process: wastewater treatment and the production of microalgal biomass, which is
of great commercial interest in agriculture and industry.
Increasing pollution of water bodies in Cartagena city and throughout the
country due to the discharge of domestic wastewater has caused an impact on the
quality of these waters, as shown by the rising concentrations of nitrogen in the
form of ammonia (NH4+), nitrites (NO2), nitrates (NO3-), and phosphorus in the
form of phosphates (PO4-2) [13]. Eutrophication of aquatic systems is caused by
an abnormal increase in their nutrients, which alter the characteristics of the food
chain and increase entropy in the ecosystem. This, results in reduced biodiversity,
where opportunistic species invade niches previously occupied by other species.
This research aims to achieve the bioremediation of phosphate compounds from
residual effluents using the microalgae Chlorella Vulgaris.
2.Materials and Method
2.1. Materials
The Chlorella Vulgaris microalga was obtained from the microalgae collection of
SENA- River and Port Nautical Center. Chlorella Vulgaris is a unicellular green
alga, its photoautotrophic growth is generally limited by lack of nitrogen, poor
lighting, pH changes, carbon limitation, etc. [14].
2.2. Cultivation conditions

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The microalgae were grown from a batch of algae. It was started with a Petri dish,
a test tube, a 250 ml Erlenmeyer and finally a 1000 ml Erlenmeyer. The strains
were kept in Conway culture medium. Culture conditions included a temperature
of 24 ± 2 °C, 39W fluorescent lamps as an artificial lighting source with
irradiation of 5000 lux, photoperiod of 12 hours of light and 12 hours of darkness,
aeration of 0.7 vvm using atmospheric air through a mechanical fan system. The
Conway medium consisted of the following components: FeCl3.6H2O,
MnCl2.4H2O, H3BO3, EDTA, NaHPO4.2H2O, NaNO3, Na2SiO, H2O, a trace metal
solution and a vitamin solution. The metal solution consisted of ZnCl2,
CoCl2.6H2O, (NH4).6Mo7O24.4H2O, CuSO2.5H2O and distilled water, the vitamin
solution was composed of Decamyl and distilled water.
2.3. Bioassays with phosphate
To analyze the phosphate concentration by spectrophotometry, a calibration curve
with standard potassium dihydrogen phosphate (KH 2PO4) at different
concentrations was first prepared. The spectrophotometer used was Genesys
brand [15,16].
For measuring the impact of Chlorella Vulgaris microalgae on the phosphate,
the microalgae kept contact with a low and high potassium acid phosphate
solution for a specific time. A sample of phosphate-free microalgae was left
acting as a reference for the bioassay. Phosphate concentration and algae growth
were measured every 24 hours for four days. Phosphate concentration was
measured by spectrophotometry at 880 nm by the ascorbic acid method and
optical density at 680 nm to quantify cell growth [17,18].
3.Results and Discussion
The microalgae were subjected to samples with K 2HPO4 concentrations of 10,000
ppm and 40,000 ppm. For the low concentration sample (10,000 ppm) in four
days of testing, a removal yield of 78.3% was achieved (see Fig. 1). Sabeti et al.
[19] used different Chlorella microalga strains for bioremediation of water with
nitrogen and phosphorus content. They reached removal of 80.9% from municipal
wastewater with an initial concentration of 212 mg/L total phosphorus. For lower
concentrations of 4 and 6 mg/L, the removal yields were estimated in 96 and
95%, respectively. A similar trend to phosphate concentration overtime was
reported by Li et al. [20], in which total phosphorus concentration decreased from
40 mg/L to 0 mg/L during the first six days.

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4 Sánchez-Aponte, J. et al.

Fig. 1. Phosphate removal low concentration sample.

For the sample with high phosphate concentration (40,000 ppm), the
microalgae removed 73.9% of the chemical added (see Fig. 2). Huang et al. [21]
evaluated the removal of phosphorus in Microbial fuel cells wastewater (MFCW)
using Chlorella Vulgaris and reported yields less than 37.22% owing to the high
phosphorus content of MFCW. Hence, it was expected that removal yield was
lower for samples with high phosphate concentration than for low concentration.

Fig. 2. Phosphate removal high concentration sample.

The optical density was measured at a wavelength of 680 nm for a week, to
demonstrate the effect of excess phosphate on the microalgae cell growth (see
Fig. 3). As is observed, the growth of the cells in the phosphate samples was
lower than the growth obtained in the phosphate-free sample. On the fifth day, the
sample containing a low phosphate concentration reached its declining stage; the

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 Treatment of high-phosphate (K2HPO4) wastewater… 5

high phosphate concentration sample reached its stationary phase, while the
phosphate-free sample was still in the growth stage. However, a considerable
increase was observed on the eighth day. Dziosa et al. [22] studied the optical
density of Chlorella sp. under phosphorus deficiency conditions and reported a
low increase in optical density until the eighth day of culture but 74.8% of initial
phosphorus content was assimilated by the microalgae.

Fig. 3. Optical density for low concentration, high concentration and

phosphate-free samples.

4.Conclusions
This work was focused on the evaluation of Chlorella Vulgaris microalgae for
removing phosphates from wastewaters in Cartagena de Indias (North Colombia).
To this end, the effect of initial concentration of phosphates in removal yields was
assessed. For the low concentration sample (10,000 ppm) in four days of testing, a
removal yield of 78.3% was achieved. For the sample with high phosphate
concentration (40,000 ppm), the microalgae removed 73.9%. Results evidenced
the growth inhibitory effect that K2HPO4 has in high concentrations on the
Chlorella Vulgaris microalgae. Still, the microalga showed a high capacity to
remove phosphate from high-phosphate wastewater.
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