Dear Alaa Cristina Al Laham thank you for posting this very interesting technical question. As an inorganic chemist I'm not a proven expert in analytical chemistry. According to the research article cited below the basic principle of the coloorimetric determination of flavonoids with aluminum chloride is the "formation of acid stable complexes with the C-4 keto group and either the C-3 or C-5 hydroxyl group of flavones and flavonols in additionwith aluminium chloride. Aluminium chloridealso forms acid labile complexes with the ortho - dihydroxyl groups inthe A- or B-ring of flavonoids." To the best of my knowledge, aluminum chloride hexaydrate gives colorless solutions in methanol, so this should not interfere with the colorimetric measurements. However, the addition of aluminum trichloride would be a deviation from the standard procedure and should better be avoided. As described in the reference, it is advisable to use a mixture of distilled water and methanol as the sample blank. Quantification of Total Flavonoid Content and antioxidant activity in comparison to a reference flavonoid as in vitro quality evaluation parameter for assessing bioactivity of biomarkers in herbal extracts or formulations

Article Quantification of Total Flavonoid Content and antioxidant ac...

This article has been posted by the authors as public full text on RG, so it can be freely downloaded as pdf file.

I hope this helps. Good luck with your work and best wishes, Frank Edelmann