2. Experimental
A digested sludge from the municipal wastewater treatment
plant in Błonie, Poland, was applied for the experiments. The pH
of the sludge was 7–7,8, alkalinity—3194–4136 mg CaCO3/dm3
and the volatile fatty acids content—444–899 mg CH3COOH/dm3
(analyses performed by wastewater treatment plant laboratory).
The process was carried out for 14 days in 1 dm3 anaerobic
bioreactors equipped with a gasometric cylinder, under nitrogen, in
32 ◦C. The stationary conditions with periodic mixing were applied.
The raw to digested sludge ratio was 36:64. The tested substances
– diesel fuel and spent engine oil respectively – were added in
amount of 10% of sludge weight (based on the concentration value
limiting the biodegradation of hydrocarbons in soil environment).
The bioreactors with the sludge without petroleum product
were used as a control.
The chemical analyses of the sludge were accomplished before
and after 14 days ofthe process, in accordance with Polish Standard
methods [18–23]:
PN–ISO 6060:2006—COD
PN–EN 16169:2012—total nitrogen (N–Kjeldahl’s)
PN–EN 14671:2007—N–NH4
+
PN–EN 26777:1999—N–NO2
−
PN–C-04576-08:1982—N–NO3
−
PN–C-04537-7:1998—orthophosphates
The amount of the biogas and its composition was determined
applying the GA 2000 PLUS analyzer.
The microbiological analyzes covered the estimation of the
number of bacteria: total mesophilic, anaerobic, acetate producing,
sulphate-reducing as well as bacteria of I–III phases of the
fermentation and methanogenic bacteria.
The total number of mesophilic bacteria was determined in
accordance with PN–EN ISO 6222:2004 [24]. The number of anaerobic
bacteria was analyzed similarly, but the plates were incubated
in ANAEROCULT anaerobic chambers. The acetate producing bacteria
were determined on the medium with ethanol [25] after
48 h of incubation in 32 ◦C and the sulphate-reducing bacteria—on
Starkey’s medium, after 7 days ofincubation. Bacteria ofI–III phases
and methanogenic were cultivated on liquid media developed by
Grabinska-Łoniewska ´ and Słomczynski ´ [26]. For the bacterial cultures
in solid microbiological media the results were presented as
cfu (colony forming unit) in 1 g of the sludge dry weight, while for
these carried out in liquid media it was calculated as the number
of the cells/g of the sludge dry weight.