HPLC-GCXGC/FID (high-performance liquid chromatography followed by comprehensive two-dimensional gas chromatography with flame-ionization detection) and GCXGC/ToF-MS (comprehensive two-dimensional gas chromatography with time-of-flight mass spectrometry) were used to study the biodegradation of petroleum hydrocarbons in soil microcosms during 20 weeks. Two soils were studied: one spiked with fresh diesel and one field sample containing weathered diesel-like oil. Nutrient amended and unamended samples were included. Total petroleum hydrocarbon (TPH) levels in spiked soil decreased from 15,000 to 7,500 mg/kg d.m. and from 12,0O0 to 4,000 mg/kg d.m. in the field soil. Linear alkanes and aromatic hydrocarbons were better biodegradable (>60% degraded) than iso-alkanes; cycloalkanes were least degradable (<40%). Aromatic hydrocarbons up to three rings showed better degradability than n-alkanes. GCXGC/ToF-MS analysis of leaching water showed that initially various oxygenated hydrocarbons were produced. Compound peaks seemed to move up and rightward in the GCXGC chromatograms, indicating that more polar and heavier compounds were formed as biodegradation proceeded. Nutrient amendment can increase TPH removal rates, but had adverse effects on ecotoxicity and leaching potential in our experiment This was explained by observed shifts in the soil microbial community. Ecotoxicity assays showed that residual TPH still inhibited cress (Lepidium sativum) seed germination, but the leaching water was no longer toxic toward luminescent bacteria (Vibrio fischeri).

In the present study evidence is given for the presence of an electrogenic, vacuolar type ATPase (V type ATPase) in the apical membrane of malpighian tubules of Formica. Barium (6 mM), the metabolic inhibitor monoiodo-acetic acid (MIA; 5 10^-4M) and two inhibitors of V type proton ATPases, i.e. bafilomycin A1 (Baf-A1; 5·10^-6M) and N-ethylmaleimide (NEM; 5 10^–4M), all inhibited fluid secretion significantly (p < 0.05). This is in agreement with the hypothesis that K enters passively via K channels in the basolateral membrane and that a V type ATPase is involved in the active transport step at the apical membrane. Also MIA, NEM and Baf-A1 slightly depolarized the apical membrane potential, V_ap, by 16, 17 and 30 mV, respectively, whereas they had virtually no effect on the basolateral membrane potential (V_m). The mild effect on V_ap, in contrast with the pronounced effect on fluid secretion, can be explained by the high apical over basolateral membrane resistance: the voltage divider ratio, VDR, was 47 ± 9 (n = 6). As a consequence the basolateral membrane will impose its value on the other barriers. VDR was decreased to 1.4 ± 0.2 (n = 19) by Ba. Ba also caused a strong and reversible hyperpolarization of both V_b1 and V_ap (from – 16 ± 1 to – 84 ± 4 (n = 8) and from -51 ± 4 to-96 ± 6 mV, respectively). As expected MIA, NEM and Baf-Al now had a much more pronounced depolarizing effect, i.e. they drastically reduced the Ba-induced hyperpolarization of both V_ap and V_b1 The reduction in V_ap was 67, 67 and 54 mV, respectively. V_b1depolarized from -73 ± 4to-15 ± 4mV (n = 7), from-72 ± 10 to 13 ± 2mV(n≈5)and from – 78 ± 3 to – 30 ± 5 mV (n = 12) in the presence of Ba and MIA, NEM or Baf-A1, respectively. From cable analysis and total transepithelial resistance in the absence and presence of barium it was also possible to make an estimate of the resistances across the different barriers: total basolateral resistance = 10 Ω·cm², total apical resistance = 475 Ω·cm², total shunt resistance = 228 Ω·cm². It was concluded that in malpighian tubules of Formica an H pump of the V type is present in the apical membrane. As suggested in other epithelia this pump can be the prime mover in active K transport: the proton concentration gradient built up across the apical membrane can drive a K/H exchanger.

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