HPLC to UHPLC Conversion Notes (Gradient Time Program Adjustment)

In an earlier article we discussed how to adjust the flow rate, injection volume and column dimensions when scaling an HPLC method UP or Down. The formula's needed to do this are fairly simple. If we adjust for changes in the column dimensions or flow rate, what types of changes are needed to adjust the gradient time? The formula to make this adjustment is also very simple. Here is the information you need.

Terms Used in Formula:

Time in minutes, Gradient (Initial): Tg1

Time in minutes, Gradient (New):   Tg2

Flow Rate in mL/min, Column (Initial): Fc1

Flow Rate in mL/min, Column (New): Fc2

Column Diameter, mm (Initial): Dc1

Column Diameter, mm (New): Dc2

Column Length, mm (Initial): Lc1

Column Length, mm (New): Lc2

• Tg2 = Tg1 x (Fc1/Fc2) x ((Dc2²) / (Dc1²)) x (Lc2 x Lc1)

Here is an example problem to solve for. 

If we start with a flow rate of 1.000 mL/min (Fc1) on a 4.6 x 250 mm column with 5 micron support (Dc1 & Lc1) and have an initial Gradient Time of 10 minutes (Tg1), then what would the new gradient time be if we switched to a sub 2 micron support in a 2.1 x 50 mm column (Dc2 & Lc2) at 0.200 mL/min (Fc2)? 

To solve the equation we will plug-in the values for each part of the equation separately, then multiply them to obtain the result.

  (Fc1/Fc2):    1.000/0.200 = 5

  (Dc2²) / (Dc1²)  4.41/21.16 = 0.21

   (Lc2 x Lc1) = 50/250 = 0.20

  Tg2 = 10 x 5 x 0.21 x 0.20 

  Tg2 = 2.10 (or 2.10 minutes)

If a 2.1 x 50 mm column was substituted for the 4.6 x 250 mm AND the flow rate was changed from 1.000 mL/min to 0.200 mL/min, then the initial programmed gradient time of 10 minutes would be changed to 2.1 minutes

Ion Exchange Resin Mesh Size Number to Millimeter (mm) Conversion Table

Chromatography Ion Exchange Resins are often sold based on Mesh Size Number. Here is a table showing the conversion to particle size in microns or millimeters.

Mesh Size / Number	Particle Size (mm)
1250	0.010
625	0.020
550	0.025
400	0.037
325	0.044
270	0.053
230	0.063
200	0.075
170	0.090
140	0.106
120	0.125
100	0.150
80	0.180
70	0.212
60	0.250
50	0.300
45	0.355
40	0.425
35	0.500
30	0.600
25	0.710
20	0.850
18	1.000
16	1.180
14	1.400
12	1.700
10	2.000
8	2.360
6	3.350
4	4.750

Vacuum Pressure Units Conversion Table

Several of the questions I receive each week by email deal with scientific calculations or conversion of various units. One popular request relates to the conversion of micrograms, ppm and percent. Several years ago to address this question, I posted a table of weight to ppm units ("Conversion Factors microgram, nanogram, ppm, ppb and percent") which has proven to be very popular.

Because of the large number of vacuum pumps attached to HPLC and MS systems, another common conversion question relates to vacuum units. Due to the different applications and regions of the world, the desired unit often varies. It is for this reason that I develop unit conversion tables as I find these tables provide for a convenient way to print out and/or keep handy in a binder for future reference. Widespread computer use coupled to freely available page reader software (e.g. Adobe PDF) provides another means to store useful information as a pdf file too. I present this "Vacuum Pressure Units Conversion Table" in a viewable and an optionally available downloadable form [click HERE to download].

VACUUM PRESSURE UNITS CONVERSION TABLE:

*Some of the more commonly used values are shown in boldface type. ** Absolute Vacuum..

% Vacuum	Torr (mm Mercury)	kPa abs	Inches of Mercury	Micron	PSI
0.0	760.0	101.4	0.00	760,000	14.7
1.3	750.0	99.9	0.42	750,000	14.5
1.9	735.6	97.7	1.02	735,600	14.2
7.9	700.0	93.5	2.32	700,000	13.5
21.0	600.0	79.9	6.32	600,000	11.6
34.0	500.0	66.7	10.22	500,000	9.7
47.0	400.0	53.2	14.22	400,000	7.7
50.0	380.0	50.8	14.92	380,000	7.3
61.0	300.0	40	18.12	300,000	5.8
74.0	200.0	26.6	22.07	200,000	3.9
87.0	100.0	13.3	25.98	100,000	1.93
88.0	90.0	12	26.38	90,000	1.74
89.5	80.0	10.7	26.77	80,000	1.55
90.8	70.0	9.3	27.16	70,000	1.35
92.1	60.0	8	27.56	60,000	1.16
93.0	51.7	6.9	27.89	51,700	1.00
93.5	50.0	6.7	27.95	50,000	0.97
94.8	40.0	5.3	28.35	40,000	0.77
96.1	30.0	4	28.74	30,000	0.58
96.6	25.4	3.4	28.92	25,400	0.49
97.4	20.0	2.7	29.14	20,000	0.39
98.7	10.0	1.3	29.53	10,000	0.193
99.0	7.6	1.0	29.62	7,600	0.147
99.87	1.0	0.13	29.88	1,000	0.01934
99.90	0.75	0.1	29.89	750	0.0145
99.99	0.10	0.013	29.916	100	0.00193
99.999	0.01	0.0013	29.9196	10	0.000193
100	0.00	0	29.92	0	0