What wavelengths are you using? Do you have any double bonds in your compounds? If so you can try 214 nm.

If there is nothing you can try an refractive index or light scattering detector.

Hello.

Options for HPLC are the following detectors:

1) Charged aerosol detector (CAD)

2) Evaporative light scattering detector (ELSD)

3) Refractive index detector (RID)

4) Conductivity detector (CD)

5) Pulsed amperometric detection (PAD)

You didn't specify what type of molecules you want to see the separation of. There is lots of chemistries that are run by the detectors above such as simple sugars, amino acids, lipids, nanoparticles, both classical polymers and biopolymers, inorganic salts. But below are some links to some of the popular vendors with proposed applications.

CAD and ELSD:

https://www.waters.com/nextgen/us/en/library/application-notes/2022/comparing-elsd-and-cad-performance-on-polysorbate-quantification-in-infliximab-drug-products.html?srsltid=AfmBOoqOTNivstaOcWIPj8_A5mGpX3bweUMUbJ3-y76SguzQI_zke9-4

CAD:

https://tools.thermofisher.com/content/sfs/posters/PN-70533-quantitation-characterization-ipids-hplc-cad.pdf

ELSD (scroll down the article for applications):

https://www.waters.com/nextgen/us/en/library/application-notes/2004/waters-2420-evaporative-light-scattering-detector.html?srsltid=AfmBOopueZxlRpD5dIfSJahigKtp1-NfyTMNAg0lhEeor6kr67MEDGTe

RID:

https://resources.perkinelmer.com/lab-solutions/resources/docs/app_sugars-in-maple-syrup.pdf

CD:

https://assets.thermofisher.com/TFS-Assets/CMD/Application-Notes/AN-185-IC-Organic-Acids-Inorganic-Anions-Boric-Power-Plant-Water-LPN1996-EN.pdf

PAD:

https://assets.thermofisher.com/TFS-Assets/CMD/Application-Notes/56256-AN179_released032707-R2.pdf

Thank you for all the responses.

• The compounds in question do not contain double bonds.
• They are expected to be organic salts, such as charged amino sugars.
• The sample is a complex, heterogeneous mixture that will require separation into its individual components for detailed analysis.
• Working on extremely small sample quantities, in the range of microgram/ml or nanogram/ml, requiring enhanced sensitivity. A suitable detector module, or technique, that can be integrated into the existing LC-MS system to replace the current PDA detector is being sought for this purpose.

Chiranjeev Sharma If you just need a secondary detection before the mass spec detector, I would think about derivatizing the sample. If amino sugars is all there is (and other other small amines), think about derivatization with FDNB or by the AccQ-Tag platform from Waters. Both ways result in added chromophore to the amine with ability to analyze by UV or DAD, or fluorescence detection, either detection able to go down to ng/mL. High resolution is achieved on standard RP columns with TFA or FA/ACN mobile phase. Check the attach literature below and think if you need to alter any of the acids/salts to make it more LC-MS friendly.

If simple sugars are also present, consider running orthogonal (additional) method by PAD (ion chromatography) detection for the simple sugars. Alternatively, if you are open to outsourcing your samples, reach out to a contract testing lab or university with capabilities to run ion-chromatography with in-line mass spec detection. That will be able to analyze both amino acids, amino sugars, and free sugars together, usually without need for derivatization.

FDNB Derivatization:

Article Validated HPLC Method for Quantification of Pregabalin in Hu...

AccQ-Tag Platform:

https://www.waters.com/nextgen/us/en/education/primers/comprehensive-guide-to-hydrolysis-and-analysis-of-amino-acids/derivatization-of-amino-acids-using-waters-accqtag-chemistry.html?srsltid=AfmBOoody4KOOaYJFH4JcMEax8T5YL5jH42OsoeTZRu0tnpU9-4XUZp_

Ion-Chromatography with mass spec:

https://www.thermofisher.com/us/en/home/industrial/mass-spectrometry/ion-chromatography-mass-spectrometry-ic-ms.html

Compounds with minimal or no UV absorption present challenges in liquid chromatography purification, necessitating alternative detection methods like refractive index, conductivity, mass spectrometry, and fluorescence detection. Advanced techniques such as near-infrared and Raman spectroscopy, along with machine learning for data analysis, enhance real-time monitoring of these compounds during chromatographic processes.