Microbial enhanced oil recovery (MEOR) is an economical and efficient tertiary recovery technology that can be used to increase the recovery of heavy oil reservoirs after steam thermal operation. However, the introduction of high-pressure steam with a temperature as high as 370 °C during thermal recovery can disrupt the indigenous microbial flora of oil reservoirs. Consequently, the effective activation of the functional microbial flora after steam thermal operation is crucial for heavy oil recovery. As such, we investigated the effects of different activation methods on oil viscosity reduction, biogas production, microbial community structure, and microbial metabolic performance. The highest viscosity reduction (61.59%), methane content (25.96%), and asphaltene degradation rates were achieved when low/high content of organic nutrients were alternately added in group L-H. The results of the FT-ICR MS analysis showed that the addition of a high content of organic nutrients promoted the degradation of N1 classes, and the degree of aromaticity of N1O2 class compounds (DBE = 10) was reduced. The analysis of the microbial community showed that function bacteria, such as Firmicutes and Synergistetes, were effectively activated by the alternate addition of nutrients, which could prevent the accumulated fatty acids and accelerate the asphaltene degradation and methane production through the syntrophic relationship between syntrophic bacteria and methanogens. Thus, the alternate addition of nutrients has potential application for enhancing heavy oil recovery by simultaneously reducing heavy oil viscosity and improving methane production.
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