CHARACTERIZATION OF EXTREMELY THERMOSTABLE ENZYMATIC BREAKERS (ALPHA-1,6-GALACTOSIDASE AND BETA-1,4-MANNANASE) FROM THE HYPERTHERMOPHILIC BACTERIUM THERMOTOGA-NEAPOLITANA-5068 FOR HYDROLYSIS OF GUAR GUM

An alpha-galactosidase and a beta-mannanase produced by the hypertherm ophilic bacterium, Thermotoga neapolitana 5068 (TN5068), separately an d together, were evaluated for their ability to hydrolyze guar gum in relation to viscosity reduction of guar-based hydraulic fracturing flu ids used in oil and gas well stimulation. In such applications, premat ure guar gum hydrolysis at lower temperatures before the fracturing pr ocess is completed is undesirable, whereas thermostability and thermoa ctivity are advantageous. Hyperthermophilic enzymes presumably possess both characteristics. The purified alpha-galactosidase was found to h ave a temperature optimum of 100-105 degrees C with a half-life of 130 minutes at 90 degrees C and 3 min at 100 degrees C, while the purifie d beta-mannanase was found to have a temperature optimum of 91 degrees C and a half-life of 13 h at this temperature and 35 min at 100 degre es C. These represent the most thermostable versions of these enzymes yet reported. At 25 degrees C, TN5068 culture supernatants, containing the two enzyme activities, reduced viscosity of a 0.7% (wt) guar gum solution by a factor of 1.4 after a 1.5-h incubation period and by a f actor of 2.4 after 5 h. This is in contrast to a viscosity reduction o f 100-fold after 1.5 h and 375-fold after 5 h for a commercial prepara tion of these enzymes from Aspergillus niger. In contrast, at 85 degre es C, the TN5068 enzymes reduced viscosity by 30-fold after 1.5 h and 100-fold after 5 h compared to a 2.5-fold reduction after 5 h for the control. The A. niger enzymes were less effective at 85 degrees C (1.6 -fold reduction after 1.5 h and a 4.2-fold reduction after 5 h), presu mably due to their thermal lability at this temperature. Furthermore, it was determined that the purified beta-mannanase alone can substanti ally reduce viscosity of guar solutions, while the alpha-galactosidase alone had limited viscosity reduction activity. However, the alpha-ga lactosidase appeared to minimize residual particulate matter when used in conjunction with the beta-mannanase. This could be the result of e xtensive hydrolysis of the alpha-1,6 linkages between mannose and gala ctose units in guar, allowing more extensive hydrolysis of the mannan chain by the beta-mannanase. The use of thermostable enzymatic breaker s from hyperthermophiles in hydraulic fracturing could be used to impr ove well stimulation and oil and gas recovery. (C) 1996 John Wiley & S ons, Inc.