ENHANCEMENT OF CONTINUOUS BIODEGRADATION OF SULPHATES AND ORGANIC POLLUTANTS BY DESULFOVIBRIO DESULFURICANS VIA BIOMASS RECIRCULATION

Krystyna Kosińska¹, Tadeusz Miśkiewicz^2
1 Institute of Environmental Protection, Division in Wroclaw, Poland
² Department of Bioprocess Engineering, Wroclaw University of Economics, Poland

ABSTRACT

Liquid manure from an industrial pig fattening plant was degraded by Desulfovibrio desulfuricans bacteria in a flow reactor with and without biomass recirculation at a hydraulic retention time of 3 to 25 days. In contrast to the reactor without biomass recirculation, the time of wastewater retention in the reactor with recirculation did not affect the efficiency of either COD or SO₄^2- removal. The extent of reduction in SO₄^2- varied from 96.2 to 98.7%, while that in COD ranged from 85.5 to 91.1%. The higher performance of the reactor with biomass recirculation may be attributed to the older age of the biomass and a lower pollutant load.

Key words: Desulfovibrio desulfuricans, sulphate reducing bacteria, biomass recirculation, liquid manure from an industrial pig fattening plant.

INTRODUCTION

Desulfovibrio desulfuricans organisms are sulphate reducing bacteria (SRB). The biochemical processes induced by these bacteria are referred to as sulphate respiration, dissimilatory sulphate reduction or biological desulphurication. Sulphate respiration is a link in the chain of microbiological transformations of sulphur compounds in the environment. In the course of the process, SRB reduce sulphates in the presence of organic compounds, as described by the following formula [22]:

2 lactate + SO₄^2- → 2 acetate + 2CO₂ + S^2-

SRB are anaerobic bacteria and their activity manifests, inter alia, in such processes as wastewater treatment, biocorrosion, bioremediation, food decay, geochemical transformations and fuel production [1]. For many years SRB have been regarded as harmful and detrimental, and have been blamed for inducing corrosion [21], generating toxic hydrogen sulphide (especially during methane fermentation [16]), deteriorating biogas quality and raising the effluent COD level [10]. However, owing to their capability of reducing sulphates, degrading organic compounds and immobilising heavy metals, SRB are now perceived as being beneficial to environmental biotechnology [11]. Those benefits can be itemised as follows: enhanced efficiencies of methane fermentation and microbiological demineralisation (reduction in the sulphates present in the water) – which is particularly important because of the increasingly frequent use of closed-loop water circulation systems – enhanced removal of sulphates from acidic effluents produced during mining operations, or from groundwater, as well as the ability to convert readily soluble, toxic oxy-anions of heavy metals into sparingly soluble forms. For example, in the presence of D. desulfuricans, easily soluble Mo(VI) or Mo(V) containing compounds in oxygenated solutions transform into low-solubility Mo(IV) sulphides [19]. The same removal pattern applies to Cr(VI), Se(VI) U(VI) [20].

The use of D. desulfuricans organisms for wastewater treatment has been the subject of many studies [2, 3, 8, 9, 12, 13, 14, 15]. Some of those have focused on establishing the parameters of the sulphate respiration process and determining their optimal values. One of the main factors decisive for the efficiency of pollutant removal by SRB is the rate of bacterial growth, which depends not only on temperature, pH and the amount of available substrates (organics and sulphates) [5] but also on the COD/SO₄^2- ratio [6] and salinity [7].

The objective of the present study (which is a continuation of the authors’ previous research on sulphate respiration) was to enhance the sulphate respiration process by the recirculation of D. desulfuricans biomass.

MATERIALS AND METHODS

Microorganisms

The experiments were carried out with Desulfovibrio desulfuricans organisms. Their origin, characterisation, as well as the method by which they were adapted to the wastewater under study, have been described elsewhere [5]. The parameters characterising an inoculum with D. desulfuricans are listed in Table 1.

Wastewater

The wastewater made use of in our study was the effluent (liquid manure) from an industrial pig fattening plant. Samples were collected after passage of the wastewater through the vibrating screens of the treatment facility located at the breeding plant. Wastewater of that type was chosen because of the remarkable benefit it offered: a productive source of organic substrates, nitrogen, phosphorus and many microelements indispensable for microorganisms growth (assimilable ammonia nitrogen and phosphate phosphorus accounting for 55% and 64% of total nitrogen and total phosphorus, respectively). Since the wastewater samples displayed a comparatively low sulphate content, they were enriched with an additional source of sulphates, the technical FeSO₄·7H₂O from ferrous metallurgy, added in such quantities that enabled the COD/SO₄^2- ratio to be kept within the range 0.7-1.5 [6]. Another major advantage of using FeSO₄ ·7H₂O was that S^2- ions were bound by Fe²⁺ ions. The major parameters of the wastewater (after FeSO₄ ·7H₂O enrichment) are shown in Table 2.

Bioreactors

The wastewater was treated in airtight flow reactors with and without biomass recirculation, of a total volume 30 l and 6 l, respectively. The temperature of the biodegradation processes was maintained at 38°C. An electric agitator provided biomass mixing in the reactors. In the treatment system with biomass recirculation the reactor was connected to a secondary settling tank (clarifier), from which the entire volume of the settled biomass was recycled (Fig.1).

Each experiment began with filling the reactor with the inoculum in amounts equal to 1/3 of the effective volume of the reactor. Thereafter the wastewater was dosed at an appropriate rate so as to provide the hydraulic retention time (HRT) defined in the experiment design. Thus, the HRT for the system without biomass recirculation amounted to 3, 5, 10, 15, 20 and 25 days, while that for the system with biomass recirculation was 3.33, 4.29, 6, 10 and 25 days. Gases produced in the reactors flew through a washer filled with 1M Zn(COO)₂ in order to bind the H₂S released during biodegradation.

Analytical methods

Both in raw and treated wastewater, sulphates were determined by the gravimetric method (following their precipitation in the form of barium sulphate), COD was established by the dichromate method and total alkalinity by the potentiometric method. Biomass content was analysed using the gravimetric method and expressed as solids content. All the samples collected for analysis were filtered through filter paper.

RESULTS AND DISCUSSION

Table 3 shows the results of the experiments without biomass recirculation. The shortening of the HRT in the reactor was concomitant with a rise in the effluent sulphate concentration from 0.055 to 1.45 g SO₄^2-·l^-1 for the longest HRT (25 days) and the shortest HRT (3 days), respectively. This rise was initially slow but then became a rapid one (Fig. 2). Variations in COD followed a similar pattern. This implies that the treatment effects attained in the reactor without biomass recirculation deteriorate with the shortening of the HRT. The removal efficiency plots for COD and SO₄^2- followed a notably different pattern when the treatment process involved recirculation of the biomass (Fig. 2).

Table 4 includes the results of that experimental series. Thus, no considerable changes were observed in the effluent concentrations of the two pollutants although the HRT was shortened more than sevenfold (from 25 to 3.3 days). This means that biomass recirculation enhanced the treatment process, as the shortening of the wastewater retention time only slightly affected (if at all) the extent of removal.

What changed the patterns of COD and SO₄^2- concentrations in the effluent was the transformations that occurred in the wastewater under the influence of the D. desulfuricans organisms. The extent of those changes (removal of the two pollutants) was assessed by relating the efficiencies of reduction in SO₄^2- and COD to the HRT. As shown by the plots in Fig. 3, removal values were much the same for the two pollutants over the entire HRT range when the treatment process involved biomass recirculation: they varied from 96.2 to 98.7 % and from 85.5 to 91.1 % for SO₄^2- and COD, respectively.

When the treatment process was carried out without biomass recirculation, there appeared changes in the removal pattern (Fig. 3). For HRT=25 days, the extent of SO₄^2- reduction (98.2%) was very similar to that obtained in the reactor with biomass recirculation (98.7%). The same holds for the efficiency of COD removal: HRT=25 days brought about a 91.1% and 88.1% reduction with and without biomass recirculation, respectively. However, in the reactor without biomass recirculation, the extent of reduction in both the pollutants decreased as the retention time was shortened. Initially, the decrease was slow, but later it began to proceed rapidly. As a result, the extent of SO₄^2- and COD reduction amounted to 53.2% and 55.7%, respectively, for HRT=3 days.

Washout of bacteria could be one of the probable causes of the poor treatment effects in the reactor without biomass recirculation. However, as it may be inferred from the data in Table 3 and the plots in Fig. 4, biomass washout did not take place in that reactor. Biomass content was almost constant (2.4 to 2.58 g·l^-1) over the entire HRT range, which suggests that the volume of the biomass leaving the reactor together with the treated wastewater was compensated by a fast reconstruction of the bacterial population. In the reactor with biomass recirculation, however, there was continuing growth of the biomass (Table 4, Fig. 4). With HRT=25 days, biomass content equalled 3.02 g·l^-1 and rose to 12.53 g·l^-1 for HRT=3.3 days.

Since the volume of the biomass in the reactor without biomass recirculation did not undergo substantial changes, there was a rise in biomass load with both SO₄^2- and COD as the rate of wastewater flow through the reactor increased (Fig. 5). The decrease of the HRT from 25 to 3 days raised the biomass load with SO₄^2- and COD from 0.0485 g SO₄^2-·g^-1·d^-1 and 0.0443 g COD·g^-1·d^-1 (HRT=25 days) to 0.434 g SO₄^2-·g^-1·d^-1 and 0.414 g COD·g^-1·d^-1 (HRT=3 days). These loads were higher compared to those observed in the reactor with biomass recirculation, which amounted to 0.035 g SO₄^2-·g^-1·d^-1 and 0.046 g COD·g^-1·d^-1 (HRT=25 days), and 0.055 g SO₄^2-·g^-1·d^-1 and 0.067 g COD·g^-1·d^-1 (HRT=3.3 days). The results imply that one of the factors which may have contributed to the higher treatment efficiency in the reactor with biomass recirculation was the much lower load of the biomass with the two pollutants; for HRT=3 days, it was by an order of magnitude lower than when no biomass recirculation was involved. Hwu et al. [4] have found that the activity of the thermophilc bacteria biomass leaving the reactor was by 50% higher than the activity of the biomass left in the reactor. Assuming that this finding may also refer to D. desulfuricans, it can be anticipated that the described phenomenon might as well have contributed to the more effective performance of the reactor with biomass recirculation.

Both methods of wastewater treatment (with and without biomass recirculation) were compared with respect to the parameter that describes the rate of SO₄^2- and COD removal per unit volume of the reactor. The results are plotted in Fig. 6. As shown by these plots, the volumetric utilisation rate for COD and SO₄^2- in the reactor with biomass recirculation remained constant within a broader HRT range than when the biomass was not recycled. This observation points out that the treatment method with biomass recirculation is more efficient and beneficial. It should, however, be noted that in this method the values of the two pollutants show a tendency to decrease as the HRT shortens. This is a rather disadvantageous phenomenon, demanding more attention in further studies.

The problem of how to enhance the efficiency of wastewater treatment with biomass recirculation has attracted the attention of many investigators [e.g. 17, 18]. However, none of their reports has referred to D. desulfuricans. This finding prompted us to attempt the study reported on in this paper. The analysis of the results obtained allows the conclusion that biomass recirculation is the right step towards the enhancement of wastewater treatment efficiency by D. desulfuricans. It was found that an advanced age of the bacteria in the biomass evidently improved the treatment effects. The improvement was particularly distinct with much shorter HRT.

CONCLUSIONS

1. Continuous wastewater treatment carried out with D. desulfuricans in a system with biomass recirculation enhanced the efficiency of the process, allowing for an effective removal of COD and SO₄^2- within an HRT range from 25 to 3.3 days. Over that HRT range, the extent of SO₄^2- and COD reduction varied from 96.2 to 98.7% and from 85.5 to 91.1%, respectively.

2. The higher efficiency of continuous wastewater treatment in the presence of D. desulfuricans and with biomass recirculation is attributable to the biomass load with pollutants (both COD and SO₄^2-), as well as to the longer time of bacteria residence in the reactor (i.e., to the older age of the biomass).

3. Biomass recirculation (more advanced age of the biomass) turned out to be the right step towards the enhancement of the continuous wastewater treatment by D. desulfuricans.

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