There's two different things: hotbars and macros.
First off, items in bags, or clickies that have to be equipped but aren't, won't work ... from a hotbar or anywhere else.
But for equipped items or inventory-usable items that aren't in a bag, if you left-click and hold it on the item it should put an icon onto your cursor, and then you can put that in a hotkey bar to use the item (or use the appropriate number key).
Some people use the secondary hotkey bars and then bind other keyboard shortcuts to them, and some go further and even hide that secondary hotbar (so they just use the keyboard shortcuts with no visual to click).
However, there is no / command for triggering an item/hotkey, which means there' no way to call those items as part of a macro (that I'm aware of).
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