LC•GC Europe - February 2001 GC connections 1

Basic GC Measurements
and Calculations
John V. Hinshaw, ChromSource, Franklin, Tennessee, USA.

Tracking a column’s performance over its lifetime is not complicated. Gas chromatography (GC) users can learn a great
deal about their columns by making several measurements and calculations from simple test chromatograms acquired
weekly or more often. This month, John Hinshaw reviews some basic GC measurements and calculations and applies them
to questions of column quality.

Analysts often need to make calculations understand derive from measurements of respond to air in a sample, but a flame
related to their chromatographic systems. retention time and peak width made from ionization detector will not. To measure the
Yet, many do not know what calculations the chromatogram. These measured and unretained peak time, analysts purposely
are appropriate, or they use incorrect calculated values and their trends over time inject an unretained substance to which the
formulas that can cause serious errors. A are excellent diagnostic tools that indicate detector responds. Among these substances
good understanding of how to measure how well a GC system functions and point are methane or butane for flame ionization
and calculate various working parameters to developing problems before they detectors, air for thermal conductivity
from an instrument and a chromatogram become serious. detectors and methylene chloride vapour for
does not require strong theoretical Unretained peak measurements: Figure 1 electron-capture detectors.
knowledge. Some high-school algebra and illustrates a simple chromatogram with The unretained peak time can change
the willingness to make a little extra effort three peaks. The first peak is the detector during programmed temperature or
are the only prerequisites for improving the response to substances that are not retained programmed pressure GC runs —
accuracy and usefulness of common in the stationary phase. These components including constant flow control of packed
chromatographic measurements. traverse the column entirely in the mobile or capillary columns — depending on the
Calculating and applying these phase, as expressed in the designation of pneumatic system and its mode of
measurements correctly facilitates their retention time (tM, in which the operation. Record the unretained peak at
instrument and method maintenance. subscripted M indicates its non-retained the initial column temperature and
Effective method transfer from instrument characteristic). If one of the non-retained pressure to verify the instrument set-up
to instrument requires a good working solutes evokes a detector response, then the and then maintain a complete description
knowledge of chromatographic chromatogram will include a peak at this of the method temperature and pneumatic
computations as well. time. A thermal conductivity detector will conditions from beginning to end.
In this “GC Connections” column, I’ll
present a number of useful measurements
and calculations, based on a test
chromatogram, that can help gas
chromatography (GC) users to better
understand the analytical process and
thereby improve control over the quality of
mmax wh
their results.

The Chromatogram
The classic elution chromatogram is a two-
dimensional representation of the amount
of solute exiting the column as a function
of elapsed time since the separation 0 tM tR1 tR2
experiment began. In GC, this plot Time
represents detector response during the
period after solute injection. Many of the Figure 1: Basic measurements from an isothermal chromatogram. mmax is the peak height at
parameters that chromatographers must its apex and wh is the peak width at half height.
2 GC connections LC•GC Europe - February 2001

GC operators should determine the unretained peak time and columns. For any column at a specific temperature with a known
examine its shape after column installation, a carrier-gas supply carrier gas, the pressure drop across the column dictates the
change or a GC method change. This step verifies that carrier gas is average carrier-gas velocity and the column flow-rate according to
flowing through the column at the desired rate. The unretained peak relationships too complex to discuss in this “GC Connections”
should have a symmetrical shape; tailing or other distortions indicate column. Chromatographers can use these calculations if they have
a problem with the column or its installation that operators should access to a computer or to a gas chromatograph with electronic
correct before proceeding. Record the unretained peak at faster pneumatic controls. Electronic pneumatic control systems for
chart speeds — or expand its area in the chromatogram on the capillary columns incorporate the column pressure–velocity–flow
computer screen — to see its shape better. relationships into the gas chromatograph, and GC users can take
For single packed-column analyses, the unretained peak shape advantage of this built-in capability. For those without electronic
helps diagnose installation or column problems, but GC methods systems, carrier-gas calculators are available on the Internet: the flow
most often specify the column flow-rate and not the unretained peak calculator at http://www.chem.agilent.com/cag/servsup/ssmain.html
time. The unretained peak time is difficult to relate to packed column is a good example.
flow-rate and pressure drop, and often it is too short for accurate To use a program such as those or the built-in capabilities of
measurement, so chromatographers should measure the flow at the electronic pneumatic systems, enter the column dimensions,
column exit and the pressure drop across the column. The carrier-gas identity, column outlet pressure conditions and — for
relationship between flow and pressure provides a convenient an external calculator — enter the column temperature. The results
diagnostic tool. After installing a new column, record the pressure from these calculations are only as good as the information that
drop and measure the flow with a flowmeter at the initial and final users input. With an electronic pneumatic system, errors in the
column temperatures. The pressure drop will be higher at elevated input column dimensions will cause the actual operating conditions
oven temperatures — make sure that it does not exceed the upper to depart considerably from the set points.
pressure limit of the pneumatics. Then check the pressure and flow Next enter the desired average carrier-gas linear velocity in the
values again at regular intervals. A significant change in the instrument system or flow calculator. The resulting column pressure is
pressure drop at either the lower or the upper temperature the pressure increment above atmospheric pressure required to
indicates changes in the column packing or a developing leak that produce the specified linear velocity through a column with the given
will require attention. Any peak-shape aberrations indicate an dimensions at the current oven temperature. An electronic system will
ongoing problem with the inlet or the column packing. adjust the pressure automatically. With an external calculator, set the
For capillary columns, the unretained peak time is an essential indicated pressure on the capillary inlet system by adjusting the
parameter that allows chromatographers to calculate the carrier- pressure regulator accordingly.
gas velocity through the column and cross-check it with the Then inject an unretained substance and record the
measured pressure drop. The GC method may specify the carrier- chromatogram. Calculate the measured average carrier-gas linear
gas velocity, the unretained peak time or both in addition to the velocity from the unretained peak time, according to Equation 1,
pressure drop. The velocity in the column is lower near the beginning and compare it with the nominal velocity entered into the
and higher at the exit, because of expansion of the compressible electronic system or calculator. The two velocities should be within
carrier gas as it moves down the column. For simplicity, most approximately 10% of each other. A larger discrepancy indicates a
chromatographers concern themselves only with the average carrier- potential problem with a carrier-gas leak, an error in the column
gas velocity along the entire column. The following formula gives the dimensions, the wrong type of carrier gas or the wrong column
average carrier-gas linear velocity (u–) in centimetres per second in outlet pressure setting.
terms of the unretained peak time (tM) in seconds and the column The column outlet pressure indicates to the pneumatic calculator
length (L) in centimetres. whether the column vents to the atmosphere or to a vacuum
system. Larger inner-diameter columns or short columns may
u–  L/tM [1] require inlet pressure less than atmospheric pressure to deliver the
set point velocity to a vacuum system; checking the required
The carrier-gas velocity affects the column’s efficiency in pressure drop before operating the column helps prevent this
resolving solute peaks. If the velocity is too high, solutes don’t mistake.
spend enough time in the column for an efficient separation; if the Wide-bore, 0.53 mm inner diameter columns present a problem
velocity is too low, solutes broaden excessively by diffusion through with pressure drop and linear velocity correlation because the
the mobile phase. A normal range of operation exists between pressure drops they require for normal operation are very low,
these two extremes that is approximately 20–40 cm/s for helium ranging from 1.5 psig helium for a 15 m column at 30 cm/s linear
and 30–60 cm/s for hydrogen carrier gases. The exact optimum velocity to 6.25 psig for a 60 m column at the same velocity.
average velocity depends on the column specifications, Mechanical pressure gauges will not indicate these low pressures
temperature profile and solute, in addition to the carrier gas. It is accurately, and even electronic systems will have trouble at the low
impossible to choose one velocity that yields the best efficiency for end of this range. Use a direct flow-controlled inlet system, not a
all solutes in a diverse multicomponent mixture. Furthermore, large split inlet, with columns shorter than 25 m in length. Calibrate the
carrier-gas velocity changes in a temperature-programmed analysis flow controller, set the desired flow-rate, and check the flow at the
affect relative peak separation as well as peak widths and may column exit. A column flow calculator program will compute the
cause adjacent peaks to be coeluted or even reverse their elution corresponding linear velocity, so it is possible to cross-check velocity
order. For these reasons, be sure to determine the average carrier- and flow for these columns.
gas linear velocity after column installation, changing carrier-gas Column dimensions: Most chromatographers take the column
tanks or making any changes to the inlet or pneumatic systems. dimensions right off the column box and assume that they are
The exact relationships between the carrier-gas pressure drop correct. In fact, this assumption is a frequent source of problems.
and the linear velocity for capillary columns provide a diagnostic The original box or specification sheet indicates the column
tool analogous to the relationship of pressure and flow for packed dimensions correctly, but after the column is removed from the box
LC•GC Europe - February 2001 GC connections 3

there is no certainty that an operator will indication to within roughly 10% of the regularly determining the retention factors
return it to the correct box. Subsequent actual column diameter. Simply adjust the of test solutes, chromatographers can
users may be surprised to find that the input column diameter until the calculated monitor the condition of their columns and
column in the box is not what the box pressure or linear velocity correlates with the can detect small stationary-phase losses or
indicates. Most columns come with a measured value. Then round the apparent shifts in apparent column polarity before
serialized identity tag attached to the diameter up or down to the closest they significantly affect peak separation
column cage for this reason. Even then, diameter available from the specific column and resolution.
the length of a capillary column becomes manufacturer and record that value. Extended column use at temperatures
shorter with each installation — sometimes The unretained peak time provides a close to a column’s maximum rated
by as much as 1 m — which quickly check not only of the column linear operating temperature can significantly
invalidates the length shown on the tag. velocity that is important for obtaining the reduce the amount of stationary phase in
Accurate column length is essential for best possible column performance but also the column. Other catastrophic events,
several computations, including the carrier- for checking the column dimensions and such as oxygen leakage or chemical stress
gas linear velocity. With electronic the electronic pneumatic system set-up. can cause a sudden loss of stationary film
pneumatic control systems, the column Retained peak measurements: The next or a change in the phase polarity.
length and other column dimensions play a items to examine in Figure 1 are the Chromatographers can track the loss of
crucial role in the instrument system’s elution times of the retained solute peaks, stationary phase by monitoring retention
determination of column flow and split designated by the subscripted R. All solutes factors of test-mixture peaks as part of
ratio, which have a direct impact on that pass through a column spend the routine column qualification. Stationary-
quantitative results. same time in the mobile phase, equal to phase losses cause peaks to be eluted at
To avoid these problems, establish an the unretained peak time. Any additional lower, earlier retention factors under the
inventory list for both packed and capillary time spent in the stationary phase is equal same temperature conditions. A peak shift
columns that correlates column serial to the difference between the peaks’ towards lower retention of more than 20%
numbers with their original dimensions and retention times (tR) and the unretained will indicate a continuing problem with
packing or coating specifications. Keep a peak times. Chromatographers call this stationary-phase loss that could affect peak
log of column activities and dates of parameter the adjusted retention time (tR), separation and resolution.
installation and removal from service. For which is measured in seconds and can be Two retained peaks: The relationship of
capillary columns, measure and record the calculated from the following formula: two peaks’ retentions also plays a major
apparent column length before installation role in chromatographic separations. Two
by counting turns and computing the tR  tR  tM [3] peaks that are close together require more
length from the following formula: resolving power from a column than peaks
The adjusted retention time is not very that are far apart. Analysts measure the
L  nd [2] significant by itself, but it plays an relative separation of two peaks in two
important role in another retention similar ways: the separation factor () and
where n is the number of column coils measurement, the retention factor (k). The the relative retention (r). These two
and d is the average coil diameter on the retention factor is the ratio of the adjusted measurements take the same values, but
column cage. Subtract the total length of retention time to the unretained peak their fields of application are different. The
column removed from both ends during time. In other words, it expresses peak separation factor is commonly used in
installation. The result is accurate enough retention in terms of multiples of the characterizing the separating power of a
for most applications: an error of one coil unretained peak time. particular column under specific analytical
on a 15 cm (6 in.) cage is equal to The formula for the retention factor is conditions, and the use of relative
approximately 0.5 m, or less than 2% of a retention appears most often in data-
t'R tR  tM
30 m column length. Do not, under any handling systems. The ratio of two
k  [4]
circumstances, remove the column from its tM tM retention factors defines both parameters:
cage, stretch it out along the laboratory
floor, and count floor tiles to determine the Interestingly, the retention factor doesn’t   k2/k1 for adjacent peaks
length. Not only will this damage the depend on the average carrier-gas linear r  ki/kst for any two peaks [5]
column by attracting dirt on the outer velocity. Increasing or decreasing the
coating, but it may damage other velocity changes both the unretained and In the first equation k1 and k2 are the
laboratory personnel as they trip over the retained peak times in proportion. This retention factors of the first and second
nearly invisible tubing. behaviour makes the retention factor very adjacent peaks, respectively, when
GC users can assume that the column useful for column characterization with a determining the separation factor. For relative
inner diameter is very close to its nominal test mixture under isothermal conditions. retention, kst defines a reference peak
value, because of the precision drawing GC operators can detect potential retention factor and ki defines the target
machines that produce fused-silica capillary problems in their separations by analyte retention factor, which could be
column tubing. In general, the tubing’s periodically monitoring the retention greater or less than the reference peak
inner diameter lies within 0.4–0.5% of factors of test substances. retention factor. The two peaks for relative
its nominal value. With the correct column A retention time shift could be caused retention calculation don’t have to be
length and carrier-gas identity, the linear by changes in the column pressure drop or adjacent.
velocity test described above will quickly by changes in the column itself, such as The separation factor is always greater
identify a mislabelled column. If the column loss of the stationary phase. Pressure-drop than or equal to 1, and it always applies to
inner diameter is unknown, a flow changes do not affect the retention factor; two adjacent peaks. It indicates the degree
calculator programme will provide an however, stationary-phase changes do. By of selectivity obtained with a particular
4 GC connections LC•GC Europe - February 2001

column and set of analytical conditions. height that one theoretical plate occupies means that the column delivers nearly all
Drifting separation factors in test mixture in the column. High performance columns the possible efficiency that it could for this
chromatograms obtained over time can produce more theoretical plates and peak.
indicate a continuing problem with column correspondingly smaller plate heights. The above calculations do not include an
polarity shifts caused by stationary-phase Narrower peak widths translate to higher adjustment for the effects of the
degradation or column contamination. performance; the relationship between the stationary-phase film on the measured
The relative retention can apply to any peak width at half-height and the plate height; they assume that no film is
two peaks in a chromatogram. It is more measured number of theoretical plates present. This is untrue, of course, because
independent of long-term changes in (Nmeas) is the column wouldn’t retain any peaks if no
column length or film thickness than phase was present, and the peaks all

( (
2
absolute retention times or retention tR would be eluted at the same unretained
factors, and it is often used in data- Nmeas  5.545 [7] peak time. The stationary-phase film
wh
handling system peak identification. broadens peaks as the analytes traverse the
Significant changes in the separation Narrower peak widths at longer retention interface between it and the mobile gas
factor or relative retention indicate shifting times have higher plate numbers. For phase and as they diffuse slowly through
column polarity in isothermal analyses. In example, a peak that is eluted at 10 min the film while inside it. The extent of this
the instance of temperature-programmed with a width at half-height of 4 s has stationary-phase band broadening depends
analyses, drifting linear velocity may also 125000 measured theoretical plates. primarily upon the film thickness. Thin-film
have this effect. Dividing the column length by the plate columns with coatings of 0.25 µm or less
Peak width: Each peak has a characteristic number yields the height equivalent to a suffer only negligible losses, but thick-film
width, which chromatographers usually single theoretical plate (H), in centimetres: columns can lose as much as 50% or more
measure at a halfway point on the peak. of the theoretical performance.
The peak width at half-height (wh), given in H  L/N [8] Rather than attempt to calculate the
seconds, characterizes the aggregate degree of efficiency loss caused by the
amount of broadening that a solute has For a 30 m column that yields the peak stationary phase, GC users should measure
undergone in its passage through the inlet, mentioned above, the measured plate the plate height or number of theoretical
column and detector. In most situations, height is 0.024 cm or 240 µm. plates when a column is new. Make sure the
the inlet and detector contributions to To gauge column performance, new column delivers close to 100% or more
peak width are negligible and the column chromatographers compute the smallest of the performance, as recorded on the
normally produces most of the broadening. possible plate height that a column could manufacturer’s test chromatogram under the
Figure 1 shows two methods for peak- deliver. The minimum theoretical plate same conditions. This level of performance
width determination. The first uses direct height (Hmin) is related to the column inner may be significantly less than the theoretical
measurement of the peak width at half- diameter (dc) and the retention factor by maximum, but it reflects the effect of the
height from the chromatogram, which only the following formula: stationary film on the plate height. If a new
requires a ruler. The second method uses column fails to yield performance similar to its
the peak area and peak height to 1
6k
11k 2 original test chromatogram, then carefully
Hmin  dc [9]
approximate the width at half-height. A 12 (1
k 2) check the column installation and the linear
triangle drawn from the peak apex to the velocity. For an old column, chromatographers
baseline through the data points at half- The value of the square-root function of should expect some performance degradation
height has an area approximately equal to k in Equation 9 ranges from 0.6 at a k over time. Replacement is mandated when
94% of the entire peak area (A). The value of 1.0 to 0.95 at a k value of 100. In the column no longer delivers a satisfactory
following equation relates the peak area this example, if the average carrier-gas chromatogram for the test mixture or
and height to the width at half-height: linear velocity in the 30 m column were analytical standard, fails to deliver a minimum
30 cm/s, then the unretained peak time plate number, or tests to a larger plate height
wh  0.94A/mmax [6] from Equation 1 would be 100 s and the than tolerated by the test procedure. Any of
retention factor from Equation 4 would be these column qualification criteria are equally
where mmax is the peak height at its apex. 5.0. At a k value of 5.0, the square-root valid; operators should choose one and apply
This formula is convenient when the function in Equation 9 is equal to 0.84. If it regularly.
chromatogram is stored in a computer, the column has an inner diameter of
because it only requires the peak area and 0.025 cm (250 µm), then the minimum Conclusion
height. Many data-handling systems include possible plate height is 0.021 cm or GC users can perform many basic
the capability to report peak widths, which 210 µm (0.84 3 0.0250 5 0.021 cm). chromatographic calculations based on a
operators should choose over the How does this theoretical value for the test chromatogram, the pressure drop
approximation shown in Equation 6. With a plate height compare with the measured across the column, its temperature and the
peak width at hand, a GC operator can value? Chromatographers can calculate the column dimensions. Making these basic
ascertain the apparent column performance per cent utilization of theoretical efficiency measurements and calculations can
level in relation to the highest performance (UTE%) to make this comparison: provide chromatographers with enhanced
the column could deliver or to laboratory quality assurance for their analytical results.
standards for column performance.
Chromatographers measure column
performance in terms of the number of
UTE%  100
( (
Hmin
Hmeas
[10]
Testing following installation and at regular
intervals will provide a record of column
performance over time that analysts can
theoretical plates (N) that the column In this example, UTE% is 100  0.021 use to gauge when to replace a column, as
delivers or in terms of the distance or 0.024, which equals 87.5%. This result well as to identify problems before they
LC•GC Europe - February 2001 GC connections 5

become serious and significantly affect

“GC Connections” editor John V. Hinshaw is