When Captain Bixby is remarking about Sam's whopper, the man on Bixby's left is scratching the back of his own head with the right hand. However, on the next immediate cut, the man is holding onto his cane with both hands. Additionally, the right hand is underneath the left.
The film first shows Mark Twain wearing his famous white suit as the author speaks to his wife Livy, while she is on her deathbed. Twain began wearing the suit only after he had finished mourning his wife's death, at which time he swore he would wear only white for the rest of his life. (Michael Shelden recounted this in the opening of his biography, "Mark Twain: Man in White -- The Grand Adventure of His Final Years.")
The epitaph quoted in the movie by Samuel Clemens was not for his wife's grave; rather, it was for his daughter Susie.
As in many biographical movies, several characters and events are fictionalized, conflated, or depicted out of order.
On the poster advertising, THE CELEBRATED JUMPING FROG, the date reads Monday, May 12, 1867. However, May 12, 1867 was a Sunday.
In Florence, the closeup shots and two shots of Livy and Mark alternate back and forth. However, in the closeups, they are fully lit, but in the two shots, they are in the dark.
In the young Sam's first steerage of a steamboat, he is told to avoid trees which are off the port (left) side, but when the scene cuts to the over-the-shoulder shot of Sam, the trees are on the starboard (right) side. He then turns the wheel to the right (starboard) which should crash the boat directly into the trees. But, when it cuts back to the full shot of the vessel, the ship is successfully turning away from the trees which again are on the port side.
(at around 1h 31 mins) When Livy tells Twain she has read Huckleberry Finn, the boom mic shadow can be seen moving across the far wall behind them.