Hi everyone!
I project to measure levels of total phenolic compounds in peppers following the method of the Folin-Ciocalteu's reagent, and I am going to calculate them by reading absorbance using gallic acid as a standard.
Nevertheless, I am confused about what wavelenght should be setted: while some authors read absorbance at 765 nm (Dogan et al., 2018; Kupina et al., 2018); others read it at 760 nm (Ghasemnezhad et al., 2011); 750 nm (Toledo-Martín et al., 2015; Lwin et al., 2022) or even 725 nm (Vega-Gálvez et al., 2009).
I would be grateful if someone could give me some light about this.
Thanks to everyone in advance.
Pablo
Literature cited:
Dogan, A.; et al. (2016). Comparison of pesticide-free and conventional production systems on postharvest quality and nutritional parameters of peppers in different storage conditions. Scientia Horticulturae 207: 104-116.
Ghasemnezhad, M.; et al. (2011). Variation in phenolic compounds, ascorbic acid and antioxidant activity of five coloured bell pepper (Capsicum annum) fruits at two different harvest times. Journal of Functional Foods 3: 44-49.
Kupina, S.; et al. (2018). Determination of total phenolic content using the Foling-C assay: Single-Laboratory validation, First Action 2017.13. Journal of AOAC International 101 (5): 1466-1472.
Lwin, H.P.; et al. (2022). Perforated modified atmosphere packaging differentially affects the fruit quality attributes and targeted major metabolites in bell pepper cultivars stored at ambient temperature. Scientia Horticulturae 301: 111131.
Toledo-Martín, E.M; et al. (2015). Application of visible/near-infrared reflectance spectroscopy for predicting internal and external quality in pepper. Journal of the Science of Food and Agriculture 96: 3114-3125.
Vega-Gálvez, A.; et al. (2009). Effect of air-drying temperature on physico-chemical properties, antioxidant capacity, colour and total phenolic content of red pepper (Capsicum annuum, L. var. Hungarian). Food Chemistry 117: 647-653.
Dear Pablo Reguera ,
In short it is basically as said here by the Sigma-Aldrich site (https://www.sigmaaldrich.com/NL/en/product/sial/f9252) “Addition of the phenol reagent generates chromogens that give increasing absorbance between 550-750nm. Normally, absorbance at the peak (750nm) or shoulder (660nm) are used to quantitate protein concentrations between 1-100 mg/ml while absorbance at 550nm is used to quantitate higher protein concentrations.”.
Or as said in one of the most cited papers when it comes to Folin–Ciocâlteu reagent “Because of the breadth of these peaks and the fact that other components in biological samples do not absorb in this region, analysis can be carried out at a wide range of wavelengths, 760 nm generally being chosen for FC.” Article Analysis of Total Phenols and Other Oxidation Substrates and...
See for what they mean by breadth for example the absorbance spectra in the following paper (Figure 1): Article Application and Analysis of the Folin Ciocalteu Method for t...
Although the overall message is that every wavelength between 750-765 nm is fine, personally, I would go for 765 nm. This is based on the official JAOAC method: https://academic.oup.com/jaoac/article/102/1/320/5658204 but as said every wavelength is fine as long as you clearly indicate in your method which wavelength you choose (and on which paper you based that choice).
Best regards.
generally wavelength go between 750 to 765 nm for polyphenolic compounds determination
Pablo Reguera, I am working in the same area and measuring TPC (Total phenolic content) at 765nm and getting good results. You can go for the same. Hope this will help you.
Regards
Preety Rohilla
From the one hand, the use of suitable solvents in the mobile/mobility phase coupled with sample preparation instructions, using the column and the detector related to the type of secondary metabolite and the degree of polarity and non-polarity of the solvent particles and the resulting bonds should be considered. On the other hand, having the standards of considered compounds helps the process of isolation and identification of compounds phenolics and flavonoids. Eventually, It should be noted that setting the appropriate temperature program and effective wavelengths (280-320) must also be applied till would be made the best result.
Regards
for the evaluation of phenolics content : total phenolic compounds (TPC). The results were given as milligram of acid Gallic per gram of dry powder (mgGAE/G.dw). 20 μL of the sample was added to 100 μL of Folin-Ciocalteu reagent in addition to 80 μL of Na2CO3 , the mixture was incubated for 10 min and the absorbance was performed at 765 nm (Atanassova.,2005).
For more details, see Sustainable Chemistry and Pharmacy 31 (2023) 100933 4 S.O. Tebbi et al.
Hi
Run scanning the wavelength ranging from 720-780 nm to check the maximum absorption intensity for your extract.
@Pablo have you solved your queries about TPC?
i am facing some errors can you please connect me
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