How do I determine the void volume in HPLC?

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The void volume of a system is usually determined by injecting an unretained standard (Uracil in RP-HPLC) that has no or very little retention on a particular phase. Slight variations in this value are explained by the extra column dead volume of your specific system configuration and set-up.

Multiply the elution time of the unretained compound by the flow rate to get the actual void volume of the system and column. To determine the column void volume alone you would need to subtract the system void volume determined without the column attached.

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