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The methylene blue sulfide test is a convenient method often used in soil microbiology to quickly detect in water the metabolic activity of sulfate reducing bacteria (SRB). In this colorimetric test, methylene blue is a product formed by the reaction and not a reagent added to the system.
The ketone of 11 is reacted with methyl lithium to yield the tertiary alcohol 12, which in turn is treated with acid to produce the tertiary cation 13. The key step of the synthesis is the π-cation cyclization of 13 in which the B-, C-, and D-rings of the steroid are simultaneously formed to produce 14 .