Why is muscle soreness worse two days after a workout than the day after?

Short answer

Because pain nerves become sensitive with a delay. NGF, which makes nerves sensitive, rose from 12 hours after exercise in rats, and in people it was raised inside the muscle 24 hours later. Soreness peaked on the second day after workouts hard enough to cause a big drop in strength; after easier workouts it peaked the next day.

Why is muscle soreness worse two days after a workout than the day after?
'Lactic acid builds up → soreness': is it true?
Muscles often hurt more a day or two after a workout than right after it. People commonly blame lactic acid buildup. We checked lactic acid, when soreness is worst, and what delays the pain, in that order.
Is it lactic acid?
The run that raised lactic acid caused no soreness
People ran for 45 minutes on flat ground and downhill, and lactic acid and soreness were measured for 72 hours. Flat running raised lactic acid but caused almost no soreness. Downhill running did not raise lactic acid, yet caused soreness. Soreness had nothing to do with lactic acid.
When does it hurt most?
24-48 hours later, and later after harder workouts
A 2022 analysis that pooled 141 leg exercise studies found soreness was worst 24-48 hours after exercise. After workouts with a smaller drop in strength it peaked at 24 hours; after workouts with the biggest drop in strength it peaked at 48 hours. In that case, soreness was still large 72-96 hours later.
Does more pain mean more damage?
Pain and damage went their separate ways
110 people did 12, 24, or 60 reps of resisting while the arm was straightened. The 60-rep group had a much larger drop in strength and damage markers than the 12-rep group, but pain when the muscle was pressed did not differ. Soreness did not reflect the size of the damage well.
So why does pain come late?
A substance that sensitizes pain nerves rose late
In rats, after exercise that resists while the muscle lengthens, nerve growth factor (NGF) rose from 12 hours later and stayed up for 2 days. Giving an antibody that blocks NGF almost completely reversed the heightened pain. In a study of 7 people doing one-leg cycling that resists the pedals, NGF inside the muscle was also raised 24 hours later.
What happens when NGF is put into human muscle?
The sore area spread widely a day later
When NGF was injected into the shin muscle of 20 people, the injection spot became sensitive after 3 hours, spread up and down after a day, and settled by day 4. Pain during movement lasted 7 days. In a study of 13 people, 24 hours after exercise, when the same irritating solution was injected, the sheath around the muscle (fascia) hurt more than the inside of the muscle.
Summary: nerves become sensitive later than the damage happens
The harder the workout, the more the peak shifted to 48 hours
Soreness had nothing to do with lactic acid and did not track the size of the damage. In rats, NGF, which sensitizes pain nerves, rose from 12 hours after exercise, and in people it was raised 24 hours later. Easier workouts hurt most the next day, and workouts with a big drop in strength hurt most on the second day.