This is based on opinion. Please don't list it on a work's trope example list.
Tear Jerker / The Book of the Dun Cow
The sacrifice of Mundo Cani, since Chauntecleer had cursed at and hated him moments before his last brave act. He even uses the dog'sCatch-Phrase out of sheer regret and misery.
Chauntecleer's sheer brokenness before the battle with Wyrm. He's completely lost hope, and it shows.
It's hard not to feel bad for Senex, especially when Cockatrice, his son, remorselessly murders him before kicking the body away. All he really wanted was respect, and he seems to almost know what will happen when it comes, making no attempt to fight back.
Lord Russel's death in The Book of Sorrows, especially since it is so slow and Chauntecleer is so desperate to save him that he feeds him as long as he can, until he realizes that he's feeding a lifeless body.
When Chauntecleer, who was been fairly optimistic about the possibility of finding and rescuing Mundo Cani, finds his body underneath the earth, having rotted into a skeleton a long time ago. Out of everything in the series, this discovery breaks him the most and in a way he never completely recovers from.
The deaths of Ferric's family despite all of John Wesley Weasel's efforts to save them.
The corruption and death of Chauntecleer, and his final moments with Pertelote. He is unable to bear the idea that Ferric Coyote forgave him for the deaths of Rachel and his pup.
The backstory of the Bird With No Tongue, who was fooled by Wyrm into drinking his essence and lost her voice to it.
Chauntecleer, hoping to rescue Mundo Cani and bring him to the surface alive, discovers that he is much too late and that the dog is dead when he finds the skeleton deep in the earth.
When he is infested with parasitic worms that add to his worst qualities, Chauntecleer becomes violently paranoid and drives away all of the animals in a rage. They don't return until the end of his life.