How do you determine the dead time experimentally in LC?

How do you determine the dead time experimentally in LC?

• A. Inject an unretained marker that travels with the mobile phase; tM equals its observed retention time.
• B. Measure the time between sample injection and first peak.
• C. Compute from column length and mobile phase velocity.
• D. Take retention time of the most retained compound.

Answer: (A)

Dead time is the time for the mobile phase to travel from the injector to the detector, including all volumes that don’t interact with the stationary phase. To determine it experimentally, inject a compound that does not interact with the stationary phase—a true unretained marker that travels with the mobile phase. The peak you observe for this marker comes out as soon as the mobile phase has traversed the system, so its retention time equals the dead time (tM). This measured tM is then used to calculate other parameters, like the retention factor for real analytes, with tR = tM(1 + k) and k = (tR − tM)/tM. In practice, uracil or another nonretained marker is often used in reversed-phase LC. Other options aren’t reliable: the time to the first peak depends on whether that peak is truly unretained; calculating from column length and velocity ignores actual system volumes; and the most retained compound’s retention time has no bearing on dead time. If the unretained marker elutes at, say, 2.0 minutes, that is the dead time.

Dead time is the time for the mobile phase to travel from the injector to the detector, including all volumes that don’t interact with the stationary phase. To determine it experimentally, inject a compound that does not interact with the stationary phase—a true unretained marker that travels with the mobile phase. The peak you observe for this marker comes out as soon as the mobile phase has traversed the system, so its retention time equals the dead time (tM). This measured tM is then used to calculate other parameters, like the retention factor for real analytes, with tR = tM(1 + k) and k = (tR − tM)/tM. In practice, uracil or another nonretained marker is often used in reversed-phase LC. Other options aren’t reliable: the time to the first peak depends on whether that peak is truly unretained; calculating from column length and velocity ignores actual system volumes; and the most retained compound’s retention time has no bearing on dead time. If the unretained marker elutes at, say, 2.0 minutes, that is the dead time.