Hello all,

I'm analyzing samples to detect monosaccharides, organic acids and alcohols in HPLC. The chromatograph consistently shows broad peaks which sometimes tend to merge together to give a very wide peak. Additionally, I've noticed that soon after detecting sugars, the chromatograph dips below the baseline to give negative peaks. I would really appreciate some advice on setting I can try changing to fix these issues. Please find the specifications of the HPLC below with settings of interest in red font.

1. HPLC: Shimadzu LC10A
2. Column: Rezex™ RHM-Monosaccharide H+ Ion Exclusion (300 x 7.8 mm)
3. Detector: RID (UV-Vis not possible)
4. Mobile Phase: 0.03 N H2SO4
5. Injection Volume: 20 uL (Direct Injection)

6. Data Acquisition Settings
- Acquisition time (Detector A)
- Sampling: 2 Hz

7. Pump Settings
- Isochratic Flow
- Flow Rate: 0.6 ml/min
- Pump Limit: 10.00 MPa (maximum)

8. Detector A
- Mode: Analytical
- Polarity: Positive
- Cell Temperature: 40 degree C
- Response: 1.5 seconds [other options: 0.05 seconds to 10 seconds]
- (Output) Intensity Unit: Volt [other option: RIU]
- Auxiliary Range: 1.0E-3 RIU/V (other options: 1.0E-4;2.5E-4;1.0E-2]
- Recorder Range: 100 uRIU/FS

9. Column Oven
- Column Temperature: 75 degree C (max. 85 C)

10. Post-run analysis (Integration)
- Width: 5 seconds
- Slope: 200 UV/min
- Drift: 0 UV/min
- T.DBL: 1000 counts
- Calculated by: Area (other option is 'Height')

Sample Preparation
- 500uL of 0.09N H2SO4
- 600uL of sample (possibly contains high dissolved CO2)
- 200uL of ISTD for organic acids (crotonic acid)
- 200uL of ISTD for alcohols (ethylene glycol)

1. Regarding negative peaks
I've recently started degassing the mobile phase using vacuum filtration along with sonication (not under vacuum) to mitigate the effects of gases in mobile phase. Additionally, my samples contain relatively high amounts of dissolved CO2, so I'm considering sonicating them too.

2. Regarding broad peaks
I suspect that this may be because the HPLC "draws" the peaks bit slowly compared it's "speed of detection". since by the time it is (slowly) completing the peak for one analyte, the next analyte has already been detected. So, instead of joining the first peak to the baseline it starts drawing the peak of second analyte - thus the joined peaks in chromatograph. I could be wrong about this though.