While some levels in Puffle Launch are likely dead, others have known improvements or are frames away from an untied. I am posting this to keep a record of which levels are the closest to being improved.
I made a mod of the game which has all levels, the timer, turbo mode and slow motion unlocked from the start, and tracks centiseconds. If you want the file ask me on the Discord server. Note that it is not valid for submitted runs.
The game rounds all times, so to cut a full second you must get below x.50. So x.54 is two frames away from untied, x.58 three frames, and so on.
Level 4: 13.58. Very possible if a balloon bounce can be cut from the ending.
Level 5: 4.54. Still can't beat 4.54 after many dupes. A different bounce may be possible.
Level 8: 8.5. I don't know if this run had any imperfections.
Level 12, 24, 36: All boss levels, not heavily optimised.
Level 13: 8.5 (not recorded).
Level 14: Bounce from the last anvil into the catapult is possible, likely saving a second.
Level 19: Very long and difficult level, under a minute is certainly doable. Panda has posted a slightly faster start.
Level 20: See the videos here. Last shot is very precise but combining the runs should save 2 seconds.
Level 21: A second faster with a single bounce on the penultimate balloon.
Level 22: Not well optimised.
Level 33: A couple of bounces can be done first try saving a few seconds.
Level 34: A few bounces on the boxes can be saved.
Level 35: Flying straight into the goal is possible and may cut a second depending on how low the decimal of the current WR was.