Bioremediation of Soils Contaminated with Oily Sludge. A Romanian Field Study

I.G. Petrisor¹, S. Dobrota¹, I. Lazar¹, A. Voicu¹, M. Stefanescu¹, M. Kuperberg²

¹ Institute of Biology of Romanian Academy, Bucharest, Romania
E-mail:ipetriso@garnet.acns.fsu.edu

² Institute for International Cooperative Environmental Research, Florida State University, Tallahassee, Florida
E-mail: mkupe@mailer.fsu.edu

Abstract

Oil extraction and processing operations in Romania have resulted in the production of large volumes of oily sludges, which constitute a severe pollution problem. The oily sludges contain crude oil (10-60%), water (30-90%) and petroleum solid particles (5-40%), in various proportions depending on their origin. These sludges are emulsions of either oil-in-water or water-in-oil type, with relative stability determined by the presence of mineral particles along with the oil and reservoir water. Previous investigations of our research group have focused on microbial communities naturally occurring in these oily sludges and have selected several bacterial strains and consortia highly efficient in degradation of sludge hydrocarbons. This paper presents the results of investigations regarding the in situ bioremediation of sludge contaminated soils by selected bacterial strains and consortia. The studies were conducted at Potlogi Oil Field, Dambovita County (southern Romania), in experimental parcels with and without addition of selected bacterial inoculum. Before inoculation, the sludge-contaminated soil was mixed with coarse sand and chopped hay in all experimental parcels except controls. These bulking agents increase the permeability and porosity of oily sludge, improving bacterial accessibility to residual hydrocarbons. A nutrient solution was added to all parcels to stimulate naturally occurring microbiota. Hydrocarbon concentrations in soil samples as well as the status of soil microbial communities were evaluated routinely. At the end of the 2-year experimental period, good hydrocarbon degradation efficiencies (exceeding 50%) were observed, being associated with rich microbial populations in the parcel soils. The best results were found in those plots to which the bacterial inoculum was added.

Introduction

Extensive oil extraction and processing in Romania have resulted in massive accumulation of oily sludges, with severe environmental impact. High volumes of sludges are formed and block the oil reservoirs, therefore being periodically removed and deposited on the neighboring land.

The oily sludges are usually stable emulsions of either water-in-oil or oil-in-water type. Consequently, the success of any sludge remediation process relies on the ability to firstly brake up the sludge emulsion, substantially reducing its volume and facilitating hydrocarbon availability to degrading agents (Castaldi and Ford, 1992; Volkering et al., 1995). Apart from the synthetic surfactants, biosurfactants are well known for their ability to brake up stable emulsions till the complete separation of the two phases. In this way, approximately 70% of the total sludge volume can be reduced, the resulting crude oil sent to refinery, and the water reused. The remaining 30% of sludge can then be subjected to bioremediation, with high chances of success. Usually, bioremediation consists either in biostimulation, involving nutrient addition and aeration to stimulate indigenous microbiota, or bioaugumentation, that also includes the addition of selected microbial inoculum (Hazen, 1997). In many cases, it is rather difficult to assess the advantages of bioaugumentation over biostimulation (Harkness et al., 1993). In addition, the higher costs involved in the first case, makes biostimulation more attractive for field applications. The effectiveness of bioremediation is influenced by a series of factors, such as the microbial adherence to substrate (Baker and Herson, 1990), and the C : N : P rate (Paul and Klark, 1989).

The present paper presents a field experiment investigating the potential of bioremediation in the cleaning up of oily sludge contaminated soils from Potlogi Oil Field, Dambovita County, in southern Romania. Both biostimulation and bioaugumentation have been evaluated in situ, on experimental parcels. The effect of coarse sand and chopped hay addition in stimulating the process has been also investigated.

Materials and Methods

The field experiment was carried out over a 2-year period (1996-1998), on experimental parcels of 2/1 m with a high of 25-35 cm. Each parcel contained oily sludge contaminated soils (with 24.66 % residual hydrocarbon content) mixed with sand, chopped hay, mineral solution (Koronelli et al., 1984) and/or selected bacterial inoculum in the following proportions:

V₁ = Oily sludge (5 %) + sand (87 %) + chopped hay (8 %)

V₂ = Oily sludge (10 %) + sand (82 %) + chopped hay (8 %)

V₃ = Oily sludge (15 %) + sand (77 %) + chopped hay (8 %)

V₄, V₅ and V₆ = same as V₁, V₂ and V₃ + bacterial inoculum (10 %)

To all parcels, 50 L of 5% mineral solution were added. The bacterial inoculum used was represented by 6 bacterial consortia, previously isolated from Potlogi oily sludge (Lazar et al., 1995 a,b) and selected for their ability to degrade different oil types and produce biosurfactants and biosolvents. The parcels were watered every 2-3 days and aerated every 15 days. For the three parcels with addition of inoculum, periodic supplementations with 10 L of inoculum and 50 L of mineral solution were performed every 30 days. These treatments were interrupted during the cold period (december-february).

The hydrocarbon degradation efficiencies, as well as the production of biosurfactants and biosolvents for the 6 bacterial consortia used as inoculum were firstly investigated in the laboratory. The hydrocarbon degradation percentages were obtained by benzene extraction procedure, using Soxhlet equipment (Bosecker et al., 1993), on three types of crude oils. The kerosene emulsifying test (Grula et al., 1983) was used to assess the production of biosurfactants, and the nigrozine test (Lazar et al., 1995 a) for the evaluation of biosolvents production. For each experimental parcel, periodic analyses of hydrocarbon degradation percentages and microbial community characteristics, including the number of aerobic heterotrophic, facultative anaerobic and hydrocarbon-oxidizing bacteria, have been performed every 6 months according to the methodology described by Lazar et al. (1995 a,b).

Results and discussions

This field experiment was planned based on the results of a series of previous lab-scale experiments (Lazar et al., 1995 a, b, 1999, Dobrota et al., 2000), conducted between 1992-1995. The selected bacterial consortia with high efficiency in degradation of sludge hydrocarbons were mainly composed by the following genera: Bacillus, Pseudomonas, Mycobacterium, Corynebacterium and Micrococcus. Six of these consortia were chosen for this field application and firstly tested in the laboratory. The results, presented in table 1, confirm their high potential for paraffinic and semi-paraffinic oil degradation and a good production of biosurfactants and biosolvents. On this basis and in accordance with literature data (Hiebert et al., 1994), it is expected that all the consortia used in this study should be efficient in degradation of hydrocarbons from oily sludges from Potlogi area. These oily sludges are rich in semi-paraffinic oil.

Table 1. Characteristics of the bacterial consortia used as inoculum in the bioremediation experiment of oily sludge from Potlogi oil field

* The intensity scale considered was arbitrary chosen between: - to +++

The results from the periodic analyses of soil samples from each experimental parcel organized at Potlogi oil field, presented in (residual hydrocarbon degradation percentages) table 2 (presence of microorganisms in Potlogi oily sludge), indicate a reduction of residual hydrocarbons of over 90%, in all the parcels, after 24 months. This is very encouraging for a field application.

As expected, the highest degradation percentages were obtained in the experimental parcels with inoculum addition, however with not significantly higher values to justify the use of inoculum. This can be explained by the good activity of the indigenous microbiota that was stimulated during this application by nutrient addition and aeration (biostimulation tests – parcels without inoculation). These results confirm the observations of Harkness et al. (1993) concerning the similar efficiency of biostimulation and bioaugumentation in sludge remediation processes.

Concerning the best proportions of sand and chopped hay, the higher rates (87% sand and 8% hay) were more efficient, but again, not significant differences in the final degradation percentages could be observed compared to the other rates used. The sand and chopped hay facilitated sludge degradation, which is in accordance with other studies (El Nawawi et al., 1992; Hiebert et al., 1994), in which these materials were reported efficient in increasing sludge permeability and porosity, along with bacterial access to pollutants. The degradation percentages increased gradually over the 2 year-period, with slightly higher rates in the first and the last 6 months.

Click image to enlarge

Table 2. The presence of microorganisms in Potlogi oily sludge, from experimental parcels at different time periods

The microbiological study of sludge samples from each experimental parcel revealed a rich microbial community, justified by the high number of heterotrophic aerobic, facultative anaerobic and hydrocarbon-oxidizing bacteria, reported over the whole 2-year period of this study. The insignificant difference in the number of bacteria from inoculated and uninoculated parcels confirm the good results obtained in degradation of sludge hydrocarbons in each case and might be due to the natural good colonization of the sludge, which makes any supplementary introduced microorganisms hard to become established and survive. 

It is of interest to mention that after the end of this 2-year experiment, bean and then corn were cultivated on all of the six parcels. The crop development was optimal, demonstrating the success of the described field experiment.

Conclusions

1.   The bioremediation experiment, carried out in situ on experimental parcels at Potlogi Oil Field in southern Romania, was successful, resulting in over 90% reduction of sludge residual hydrocarbons after only 2 years.

2.   Good biodegradation percentages were obtained in both bioaugumentation and biostimulation techniques (inoculated and uninoculated parcels), due to the existence of an already rich indigenous microbiota in the sludge contaminated soil.

3.   The addition of sand and chopped hay to sludge contaminated soil facilitated sludge hydrocarbons biodegradation, regardless to the proportion in which they were used.

References

1.   Baker, K.H., Herson, D.S. (1990) – In situ bioremediation of contaminated aquifers and subsurface soils. Geomicrobiol. J. 8, 133-146.

2.   Bosecker, K., Hollerbach, A., Kassner, H., Teschner, M., Wehner, H. (1993) – Chemical and microbiological studies on petroleum contaminated soils after two years of biotreatment. In: Biohydrometallurgical Technologies, Vol. II, Ed. by A. E. Torma, M. L. Apel, C. L. Brierley, 365-373.

3.   Castaldi, F.J., Ford, D.L. (1992) – Slurry bioremediation of petrochemical waste sludges, Wat. Sci. Tech. 25 (3): 207-212.

4.   Dobrota, S., Lazar, I., Voicu, A., Stefanescu, M., Petrisor, I. G. (2000) – The use of microbiological method in bioremediation of sludges from petroleum industry. In: Proceedings volume of the Nat. Conf. of Biotechnology and Environmental Engineering, Targoviste, 167-173.

5.   El Nawawi, A. S., Bagouri, I. H., Abdal, M., Khalafaouri, U. S. (1992) – Biodegradation of oily sludge in Kuwait. World Journal of Microbiology and Biotechnology 8 (6): 618-620.

6.   Grula, E. A., Russel, H. H., Bryant, D., Kenaga, M., Hart, M. (1983) – Isolation and screening of Clostridia for possible use in microbially enhanced oil recovery. In:  Proceedings of the 1982 Int. Conf. on MEOR, Ed. by E. C. Donaldson and J. B. Clark, Shangri-La, Afton, Oklahoma, 43-47.

7.   Harkness, M.R., McDermott, J.B., Abramowicz, D.A., Salvo, J. J., Flanagan, W. P., Stephens, M. I., Mondello, F. J., May, R. J., Lobos, J. H., Carroll, K. M., Brennan, M. J., Bracco, A. A., Fish, K. M., Wamer, G. L., Wilson, P. R., Dietrich, D. K., Lin, D. T., Margan, C. B., Gately, W. L. (1993) – In situ stimulation of aerobic PCB biodegradation in Hudson River sediments, Science 259, 503-507.

8.   Hazen, T. C. (1997) – The microbiology of the terrestrial deep subsurface, Bioremediation Chapter, ICRC Press, 247-266.

9.   Hiebert, F. K., Portwood, J. H., Portwood, J. T., Petersen, F. S. (1994) – On site bioaugumentation treatment of petroleum tank bottom wastes: a case study. In: Microbial Enhancement of Oil Recovery – Recent advances, Ed. by E. T. Premuzic and A. Woodhead, Elsevier, 349-354.

10.  Koronelli, T. B., Komarova, T. I., Ignatchenko, A. V. (1984) – Interaction of Pseudomonas and Mycobacterium in a mixed culture and hydrocarbon oxidation. Microbiology 53 (2): 213-217.

11.  Lazar, I., Voicu, A., Dobrota, S., Stefanescu, M., Sandulescu, L., Archir, G., Lazar, I.G., Mucenica, D., Balalia, A. (1995 a) – Microbial communities of soils, slops and waters contaminated with waste oil and phenols and their role in bioremediation of such environments. In: Biohydrometallurgical Processing, Vol. II, Ed. by C. A. Jerez, T. Vargas, H. Toledo and J. V. Wiertz, University of Chile, 365-391.

12.  Lazar, I., Voicu, A., Dobrota, S., Stefanescu, M., Sandulescu, L., Archir, G., Lazar, I. G., Mucenica, D., Balalia, A., Nicolescu, C. (1995 b) – Investigation on potential bacteria for the bioremediation treatment of environments contaminated with hydrocarbons. In: Proceedings Volume of the Fifth Int. Conf. on MEOR and Related Biotechnology for Solving Environmental Problems, Sept. 11-14, 1995, Dallas, Texas, 535-547.

13. Lazar, I., Dobrota, S., Voicu, A., Stefanescu, M., Sandulescu, L., Petrisor, I.G. (1999) – Microbial degradation of waste hydrocarbons in oily sludge from some Romanian Oil Fields. Journal of Petroleum Science & Engineering, Elsevier, 22, 151-160.

14.  Paul, F. A., Klark, F. G. (1989) – Soil Microbiology and Biochemistry, Academic Press, San Diego.

15.  Volkering, F., Breure, A. M., Van Andel, J. G., Rulkens, W. H. (1995) – Influence of nonionic surfactants on bioavailability and biodegradation of polycyclic aromatic hydrocarbons. Appl. Environ. Microbiol. 61 (5): 1699-1705.

Top