Dear friend Yasuhiro Nishida

Ah, ELSD, the Evaporative Light Scattering Detector, a gem in the world of HPLC detectors! Now, let me share some insights.

Firstly, ELSD is often a savior when dealing with compounds that lack UV absorption or those that don't have a chromophore. It detects analytes based on their ability to scatter light when eluted from the column. Now, let's delve into the real-world scenarios:

**Advantages:**

1. **Universal Detection:** One of the main advantages is its universality. It can detect virtually any compound regardless of its optical properties, making it a fantastic choice for compounds with no UV absorption.

2. **Quantification of Analogous Compounds:** ELSD is particularly useful when dealing with structurally analogous compounds that might not have distinct standards. This makes it valuable for natural product analysis or in cases where obtaining pure standards is challenging.

3. **Low Detection Limits:** ELSD often provides lower detection limits compared to other detectors, which is beneficial when dealing with trace-level analysis.

**Concerns:**

1. **Baseline Drift:** ELSD is known for baseline drift, which might complicate the quantification of compounds. Strategies like using an internal standard or appropriate calibration techniques are often employed to address this issue.

2. **Sensitivity to Mobile Phase Changes:** Variations in the mobile phase composition can affect the signal intensity. Users need to carefully optimize the mobile phase to get consistent results.

3. **Sample Dependent Sensitivity:** The sensitivity of ELSD can be sample-dependent, and it might require method adjustments for different compound classes.

**Review Articles:**

1. **"Lecoeur, M., Decaudin, B., Guillotin, Y., Sautou, V., Vaccher, C., & ARMED Study Group. (2015). Comparison of high-performance liquid chromatography and supercritical fluid chromatography using evaporative light scattering detection for the determination of plasticizers in medical devices. Journal of Chromatography A, 1417, 104-115. provides a comprehensive overview.

2. **"Megoulas, N. C., & Koupparis, M. A. (2005). Twenty years of evaporative light scattering detection. Critical reviews in analytical chemistry, 35(4), 301-316., is another valuable resource.

Remember, my eager interlocutor Yasuhiro Nishida, ELSD is a versatile tool, but like any technique, it has its nuances. The key is in understanding those nuances and wielding them to your Yasuhiro Nishida advantage in the quest for chromatographic mastery!

Dear Yasuhiro Nishida,

I have attached herewith research paper for your reference.

The ELSD is NOT a "Universal" detector (Unscientific misinformation from sales people and/or found in unscientific articles, brochures etc). Universal, far from it, LOL! *Mobile phase volatility and sample stability will have a huge effect on signal levels. The selected ELSD instrument and "settings" used for the method will result in different results. ELSD and/or CAD are for very experienced users and require a lot of setup, optimization, tuning, re-calibrating and cleaning. **Output is rarely linear, often polynomial or requiring some other type of custom curve fit, over each order of magnitude (make sure your CDS software can deal with such complex curve fitting for EACH sample!). Each sample's response factor can vary by as much as an order of magnitude across a gradient. We never recommend them for students or users with less than ten years of chromatography experience. General recommendation to most clients is: Do NOT listen to biased sales people selling them. Do not use one. Instead, find someone with a LC-MS system and a great deal of method development knowledge to help you. This is the best general advice.

• I would like to stress that ELSD or CAD detectors should only be considered IF: (1) Sample is non-volatile and No other detection system is available for your sample and...(2) only consider using IF you are a very experienced chromatographer with many years of professional full-time experience (pref in industry) developing methods for many different types of samples. Using an ELSD is nothing like setting up a UV/VIS or RID.

These detectors have many of the same site prep issues as a LC-MS system. They require large volumes of high purity gas (N) to operate. They exhaust out the same large volume of gas (direct to a water trap, then into a suitable fume hood). Use only volatile buffers, solvents and/or additives (just as with a LC-MS). All of the samples that you run through them get baked onto the inside of the detector (drift tube and nebulizer). This material regularly contaminates the light chamber as it breaks off resulting in noisy signals. Regular cleaning (disassembly) is required to maintain them and when you put the detector back together you usually get different results. Reproducibility of methods is one of the many issues people have using them (The sales people rarely mention these and many users are unaware of them). You will need to optimize the temperature(s) and gas flow rates for every sample, in every method, every time you use the detector (very time consuming and challenging when you run a calibration). Poor for QC testing.

As a scientist who was directly involved with their designs and commercialization many decades ago (ELSD's go back to the 1980's. I developed hundreds of new methods using them over the many years) I can say with great confidence to avoid them unless the two items I mentioned above apply. In some niche areas they can be very useful indeed for R&D, esp when running gradients as they often show stable FLAT baselines (a trademark of the ELSD). To learn more about ELSD's, just look at some of my past published journal and technical articles. I have been consulting on ELSD and CAD systems since 1990.

Note: Please learn how to research your own questions (which you could quickly do on the web using a keyword search BEFORE posting here). Learning HOW to research a question on your own is one of the most important skills you can learn. Next, be careful when asking for advice in this area as a LOT of misinformation will be found (see above answers).

Dear Dr. Kaushik Shandilya,

Many thanks for the detailed information. I understand the basic instructions and characteristics from the HPLC manufacturer, but I still would have liked to know what it is like to use it in the real world. I have other questions, so please allow me to indicate them again below.

Dear Dr. Vijay Arjun Bagul,

Thank you very much for providing the reference.

I would like to see the details.

Thank you very much again.

Dear Dr. William Letter,

I appreciate the very detailed, very clear and realistic explanation. I would like a realistic explanation like your opinion. I have used many LC detectors, including a number of mass spectrometers, but this detector is the only one I have never used. I have other questions, so please allow me to indicate them again below.

I now understand the characteristics of this detector much better, thanks to all of you. I also now understand the need for a polynomial approximation of the calibration curve. Perhaps I can solve the problem of instrument stabilization and cleaning based on my experience with several MS/MS troubles.

A big question: How different is the detection sensitivity of analogues by ELSD? Can similar compounds be considered to have the same detection sensitivity?

Article The determination of lycopene Z‐isomer absorption coefficien...

One paper uses carotenoids as an example. In order to obtain the respective absorption coefficients of analogous substances (in this case, geometric isomers with different UV maximum absorption), the authors proposed to estimate the respective absorption coefficients of each isomer in combination with a PDA detector based on a certain isomer of an existing standard, assuming that the detection sensitivity of each isomer in ELSD is the same.

Although the analogues have similar basic physicochemical properties, they are unlikely to be exact matches. If the absorption coefficients of analogues can be determined in this way, conventional analyses with technical barriers, such as obtaining sensitivity coefficients for isomers by quantitative NMR based on isomer-specific chemical shifts, may become unnecessary.

ELSD is very attractive because once sensitivity coefficients and absorption coefficients are determined, existing more stable and available reference standards and detection methods (e.g., UV/VIS detectors) can be used to more accurately analyze analogous substances.

I appreciate it.

Yasuhiro Nishida

Yasuhiro Nishida I note that you work in the chemical industry, where relatively few analytical methods are published, and not all employers provide facilities for searching and keeping up with the academic literature.

I once used ELSD for an unpublished specific project when working in industrial pharmaceutical development, a field where we did have access to the literature.

Between about 1989 and 2000 we also had MS with both GC and a particle beam detector. The particle beam detector was of little academic interest and it lacked the necessary sensitivity for the project in question; some analytes were slightly volatile and presumably they evaporated in the high vacuum. As with ELSD, the analytes have to generate the right kind of particles and that is unpredictable.

However, particle beam was widely used in industry because, when it did work, usually to identify a drug impurity, it saved a lot of development time. Unfortunately, it used too much helium. We didn't have a moving belt interface, which met similar needs.

Current LC-MS instruments are not a satisfactory replacement because they don't provide electron and chemical ionisation, which are needed for library searching, interpretation, and spectral fingerprints.

Yasuhiro Nishida asked;

• "How different is the detection sensitivity of analogues by ELSD? Can similar compounds be considered to have the same detection sensitivity?" - - What kind of "analogue" ? Structural? The answer is... we would never assume or expect that any "analogues" would have similar responses at all by ELSD. By definition they are different, not the same. Small differences in solubility, structure or orientation may and often do change response. The mobile phase you use, the various settings of the detector etc, all change response (each detector is different too). In fact, the only way you could ever know the true answer (for your compounds) is to actually run a full calibration panel of them, individually by ELSD, using the same analysis method. And that result would only relate to the specific analysis conditions used (method) and compound(s) tested for your instrument on that day. Change any of those items and the response may change too (this in not UV/VIS). When gradient analysis methods are used, response, even for the same compound (!) may be different at different retention times! ELSD response for each compound may change from semi-linear, quadratic, polynomial over one or more orders of magnitude.

ELSD and CAD systems are unlike ANYTHING you have used before. Decades of use continue to reveal that reproducibility and method transfer are such large problems with these detectors, that they are not used for most sample types.

• "Evaporative HPLC Detectors; CAD (Charged Aerosol Detector) and ELSD (Evaporative Light-Scattering Detector)"; https://hplctips.blogspot.com/2017/12/evaporative-hplc-detectors-cad-charged.html

We are using ELSD detector from last 7 years and we had found very interesting and surprising notes for ELSD.

In summary we know when to use and when never use ELSD.

Application.

1. It will helpful for UV inactive detections

Drawback

1. Non-reproducer response

2. Not working in Lipid molecules, we had carried out one data with ELSD and same compound with CAD and it have a lot difference in terms of sensitivity of the impurities

3. Cant relay on ELSD in terms of quantification

You can contact me if you want to go for in-detail discussion. But in one line conclusion, go with CAD instead of ELSD.

Dear Dr. Christopher R. Lee,

Thank you so much for your valuable comments based on your experience.

I am now working mainly on the evaluation of bioactivity and pharmacological efficacy of natural products, but I am trying to establish analytical methods when necessary. Your comment that it could be useful in some cases was very helpful.

Currently, I would like to identify and quantify impurities and analogous substances using various detectors, so I would like to cooperate with outside parties and others to promote development of analysis.

Dear Dr. William Letter,

I appreciate your very valuable guidance.

Your superior knowledge of ELSDs is very informative.

We now understand that ELSD is not well suited for the method we intend to use.

We will run a pilot operation at an outside facility to see if we can actually use it for the method that meets our objectives.

Again, thank you very much.

Dear Pranav Pandya,

I appreciate your very kind offer.

Once we have a preliminary evaluation of the detection response in the ELSDs available to us, we would be happy to contact you if you would allow us to do so.

Thank you in advance for your continued support.