UPLC - cannot achieve >=250 for signal-to-noise

[annalls26]

I am running a UPLC method and was unable to achieve >=250 for signal-to-noise for one of my peak of interest. I am setting the sampling rate at 20points/sec and Fast for filter time constant.

Does anyone encounter poor signal-to-noise while running Acquity UPLC?

[unmgvar]

you will need to tell us your mobile phase, the WL you are working at
how do you do the S/N calculation?
the main issue is the noise setting
how do you define your noise?
from which run, what is the time range? how many peak widths?

[annalls26]

My instrument parameters are detail as follow:

Column Waters Acquity BEH Phenyl, 1.7μm, 2.1x100 mm or equivalent
Mobile Phase A Acetonitrile
Mobile Phase B 0.1% Phosphate Buffer, pH 2.5
Flow Rate 0.5 mL/min
Gradient Establish a gradient with the following profile:

Time % A % B
0.0 32 68
16.5 37.5 62.5
22.0 75 25
27.0 75 25
27.1 32 68
31.0 32 68

Column Temperature:45 °C
Temperature Sample manager:10 °C
Injection Volume:35 uL
Sample loop made Partial Loop with needle overfill
Detection Wavelength:UV at 275 nm
Total Run Time:At least 31 minutes
Injector washing fluid - Weak solvent: H2O/ACN 70/30 (1200 µl)
Strong solvent: H2O/ACN 30/70 (400 µl)

Sampling rate: 20points/sec
Filter Time Constant: Fast 0.2sec
S/N calculation: Signal x 2 / Noise

Currently, using Empower 2 Ep S/N to calculate the signal-to-noise ratio.


I am checking the signal-to-noise for one of my peak at 12mins in a resolution solution.

I am unable to post the chromatogram profile.

[Mattias]

I have had a lot of problems with Acquity and noise.... I would try these things (in order):

1. Exchange the flow cell if you can borrow one from another system. If not, clean the flow cell with 30% nitric acid.

2. Premix the mobile phases as far as possible. Acquity has really bad mixing, but it shouldn't matter at 275 nm and phosphate buffer.

3. Are you sure that the noise is higher - it is not the peak height that is lower? I would change the weak needle wash to something weaker (e.g. 10% acetonitrile in water). The weak needle wash is actually injected with your sample, and it can cause band broadening if it is too strong.

[HPLCaddict]

As unmvgar already asked, what are your peak widths? If peaks are not too narrow, you might consider playing with your detector settings. Choosing a higher response time aka filter time constant might smooth the baseline considerably...