Mr. Sharma,

For metabolomics (and not only for metabolomics) the method of choice is mass spectrometry (MS); however, using our more recently developed quantitative method and model formulas for processing of the experimental MS outcome ''intensity.'' In this context, please, consider my answer to discussion [A] shown, below. Please, consider, as well as, the reference section, therein; in addition to, the references cited in our publications listed in [A]. The advantages of the methods of mass spectrometry over all currently available analytical instrumentation are explicitly emphasized.

The NMR is, no doubt, a robust analytical instrumentation, but; I underline, herein, explicitly, for structural molecular analysis. It is completely uncompetitive with respect to the methods of mass spectrometry for quantitative analysis, due to the following major drawbacks, amongst others: (i) It shows high concentration LODs of analytes; (ii) inapplicability for analyzes of complex multicomponent mixtures of analytes, due to strong overlapping effects of the NMR signals; and (iii) significant error contribution to the analytical information from the available methods for quantitative processing of the NMR experimental outcomes, respectively.

Despite, the fact that our mass spectrometric method shown briefly in [A] provides exact 3D structural analysis of the analytes, as well. At this point, please consider reference [2] in my answer to discussion [B].

In other words, within the framework of our stochastic dynamic theory and model formulas [A,B], the capability of the methods of mass spectrometry of not only exact (or abolute) quantifycation of analytes, but also of exact 3D structural analysis of the molecules is extended dramatically far beyond its current application to the analytical practice as a chiefly method for quantitative analysis. The capability of the mass spectrometry, thus, becomes several orders of magnitude higher that this one of the NMR as analytical instrumentation for qualitative, quantitative and 3D structural analyses, respectively.

Therefore, It should be understood that the employment of NMR for quantitative analysis, especially of microcomponents and traces, is a highly speculative task.

[A] [https://www.researchgate.net/post/How_can_we_detect_VOCs_in_exhaled_breath_sample_using_ISQ_7000_single_quadrupole_GC-MS_system]

[B]

[https://www.researchgate.net/post/How_to_analyze_two_different_time_point_metabolomics_data]

Dear Vishal Sharma, generic binning tools are available in many statistic packs and most NMR software packages can also output their spectral data in generic formats like CSV. And of course is qNMR a very suitable technique for quantification especially when it is targeted. NMR is also capable of analyzing very complex multicomponent mixtures. Despite the high LOD there are even commercially available qNMR products out there. So I am not going to join bashing other analytical techniques. They all have their pros and cons and it depends on the problem and many other factors, which one is the best fit for purpose. Take care...

Mr. Niemitz,

Could you specify in mode detail the instrumental method performances of qNMR? Could you provide: References; protocols; experimental outcome data of multicomponent mixtures? There are needed the following data: Resolution; LODs; LOQs; precision; accuracy; selectivity; sensitivity; recovery; repeatability; reproducibility?

In particular there is important to specify what true value is obtained: Academic true value or legislation true value?

In conditions of market economy and free market there has everything as analytical instrumentation, currently. However, when public funds (budget) are used in public Universities (Research Institutions) for the purpose of misleading the Society and assimilation of public budget via so-called ''projects'' and academic positions of the ''Anaytical Sciences'', the ''last word'' (the decision) belongs to the respective Legislative Institutions.

(From 2020 the Prosecution of the European Union is investigating projects with a nominal more than 11 million euros.)

The largest NMR manufacturers are selling both qNMR hard- and software products and I am sure they all are fulfilling the legal requirements you are referring to. Regarding your questions please read the literature and let me know if you still have any questions.

Data binning is mandatory, provided to compare among the group of samples.

Dear Vishal,

forgetting the MS versus NMR discussion for the moment (which is, as Matthias pointed out "unpleasant" and will not help you, as you have now the NMR data) and coming back to your original question:

In NMR binning is recommended, because of small changes of signal positions induced by matrix effects (pH, salt, interaction between compounds...). Doing binning in Topspin is actually rather simple. You only need to create a list of integral regions (e.g. in excel exported as plain text) to be placed in the subdirector of the topspin installation (typically ../exp/stan/nmr/lists/intrng).

The format of such a file is straight forward and looks like this:

A 1.0 #regions in PPM

# low field high field bias slope

10.0 9.96 0.0 0.0 # for region 1

9.96 9.92 0.0 0.0 # for region 2

9.92 9.88 0.0 0.0 # for region 3

...

After a 2-line header each integral region is defined by a left and right border followed by parameters related to manual bias and slope correction.

In the example above I created three adjacent regions with a typical width of 0.04 ppm.

Linewise Expansion of this file down to e.g. 0 ppm by use of Excel is trivial.

Import of a file "xyz" can be done with the command "rmisc intrng xyz".

Application of this file to a set of spectra is available in topspin as the multi_integ automation (by typing "xau multi_integ"). This automation creates a table with all number for further analysis.

Alternatively we have been using the Amix software available at Bruker which may be to my remembering free of charge for academic institutions.

I hope this helps!

Good luck

Alfred

Thank you all of you for your suggestions. Currently, I'm doing binning/bucketing of NMR data and this is necessary. I'm using Amix software for this.

Best

Regards,

Vishal Sharma