Greetings everyone. I have a question regarding protein-polysaccharide complexes from plants. Recently, I heated plants extractions in NaOH and NaCl. During heating process, the colour turns from green to brown. Does it mean that mallard reaction occurred and protein-polysaccharide complexes has been formed? How do I have to confirm that the protein-polysaccharide has been formed. I have conducted Bradford assay to measure total protein and phenol-sulphuric acid test to measure the polysaccharide content. Both test shows high content of protein and polysaccharide. Pls help me to clarify my question. Thank you very much.
Mentioned tests do not reflect the specific conjugation between carbohydrates and protein.
If you have a Mass spec or HPLC or at least a UV spectrophotometer you may follow some Maillard-derived advanced and intermediate glycation end products. Many papers indicate the main markers of the Maillard reaction such as Furosine, CML, HMF (any increasing furan-based derivates), Pyralline, pyridosine, etc...
But for your case, I think browning is observed due to the alkaline environment and induced autooxidation. Phenolic substances probably oxidized in alkaline pH and the UV absorption wavelength of the phenolic molecules shifted upwards. Try to conduct an assay in neutral to acidic conditions to validate this hypothesis.
Another reason could be the polyphenol oxidase in plant extract. This enzyme can alter the protein structure and modifications occur which result in a darkened color view. You may exclude this by using polyvinylpyrrolidone in the extraction buffer.
Thank you very much Dr Ismail for the explanation .. I will definitely conduct the analysis based on your suggestion dr.
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