Ache from electrophorus electricus electric eel type 6 s

Choline chloride (99% purity), acetylcholine chloride (≥99% purity), pyridostigmine bromide, acetylcholinesterase (AChE) from Electrophorus electricus (electric eel, Type VI-S, lyophilized powder, 200-1000 units per mg protein) were purchased from Sigma-Aldrich (Saint Louis, USA). Human butyrylcholinesterase (BuChE) was highly purified from human plasma as described. 52 K 4 [{Re 6 S 8 }(OH) 6 ]•8H 2 O was synthesized and purified according to a previously published procedure. 53 Sample preparation for luminescence detection of ACh hydrolysis. A stock solution of AChE (C = 10 -6 M) was prepared by vortex-assisted dissolution of the lyophilized enzyme in NaCl (100 mM). Stock solution of BuChE 6.7 × 10 -7 M was prepared in 50 mM phosphate buffer pH 8.0. The solutions were stored at 8 °C for no more than 2-3 days before use.
The mixture of K 4 [{Re 6 S 8 }(OH) 6 ]•8H 2 O (0.0075 mM), NaCl (1 mM) and AChE ( pH 8.4) was incubated for 5 minutes at 25 °C. Then ACh was added, and its hydrolysis immediately monitored through cluster-centered luminescence. The AChE concentration was varied in the range 10 -9 -2 × 10 -8 M. The ACh concentration was varied from 0.06 to 0.9 mM. The luminescence spectrum kinetics was recorded at 25 °C every minute within 10 min.