Hey Caitlyn It's difficult to say exactly when Macduff starts to suspect Macbeth. I would guess he knows quite earlier on. The problem is he doesn't say anything overtly.
When he's talking to Ross, the fact that he says he won't go to Scone to see Macbeth crowned seems as though maybe he's suspicious, but keeping it under his hat. "No, cousin, I'll to Fife." After that, he doesn't appear again until Act Four, by which time he's already gone to England to hook up with Malcolm and bring an army back to Scotland.
The problem with Macduff is, unlike many of Shakespeare's characters, he's not much of a talker - we certainly don't get any soliloquies from him. So, it's hard to know what is going on in his head. However, because he raises the question over why Macbeth so quickly killed the grooms (who Mac and Lady M framed for Duncan's murder), - "Wherefore did you so?" - I would say his suspicions are piqued quite early.
Hope that helps a bit!
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