TOL (Toledo, OH)- DTW 49 miles, discontinued a few years ago after years of service
FRA-EWR-JFK, EWR was a diversion. No local EWR-JFK traffic.
IAH (Houston) helicopter service to several helipads. No longer in service.
PHL-JFK on TWA in the 80's and 90's - 727 service
DCA-PHL, which isn't that short
There are a number of flights that are about 100 miles including MKE-ORD (which I haven't been aboard), CAE-CLT, LIT-MEM, SMF-SFO, etc. Some of these are primarily for connections.
Another question might be "What is the shortest flight you've taken where you were O&D traffic and not connecting?" Mine was probably 100 miles on a mainline within the US (didn't want to drive late at night), about 25 miles outside the US (between islands and quicker than a ferry).