I was preparing a phosphate buffer and tried to add glucose. This was to prepare a buffer with an energy source for a bacterial study. After autoclaving, there was a distinct change in color (yellow) which may suggest transformation of my glucose. Would that affect my experiment in anyway?
I've read that typically, glucose solutions are sterilized by filtering and it is just added after autoclaving. But certain growth mediums contain glucose which would then be autoclaved. I just got a bit confused regarding this matter.
I also found a similar thread regarding this.
The change of color of Glucose solution is a charecteristic feature of any sugar exposed to high temperatures - say, common sugar solution turning brownish after heating on stove. This is known as charring of sugars which changes their cheemical structures. So, if you are planning to use glucose for any kind of inductions, add the filter sterillized solution of Glucose after autoclaving the rest of the media components. It might not matter if you are using glucose just as a carbon source, like in YPD.
Yes i observed that the glucose is changing it's color after autoclaving! I dont think its good so we are adding it after autoclaving and we filter the solution after.
Autoclaving glucose with other nutrient solutions enhances its degradation and release of toxic substances. That is there was colour change. You can autoclave it separately or just filter the solution which will give you sterility equivalent to autoclaving. But remember to check your PH after filtration.
The change of color of Glucose solution is a charecteristic feature of any sugar exposed to high temperatures - say, common sugar solution turning brownish after heating on stove. This is known as charring of sugars which changes their cheemical structures. So, if you are planning to use glucose for any kind of inductions, add the filter sterillized solution of Glucose after autoclaving the rest of the media components. It might not matter if you are using glucose just as a carbon source, like in YPD.
It is always advice not to autoclave glucose in phosphate buffer. You autoclave your Phosphate buffer and wait to cool down to room temperature. Then in separately pass your glucose solution in a sterile container through a sterile membrane filter and add the required quantity in a hood in sterile condition, mix gentler and use.
The change in colour is actually due to the formation of carbon. The best way to sterilize any solution is through filteration (0.2 micro filter) and this must be done in a sterilized environment (CCH). One of the conditions that must be put in consideration is the composition of the solution you want to sterilize. If the solution contains compounds that react or change with high temperature like that of autoclave sterilizer, it is reasonable not to use it. A good example is the cell culture medium which contains vitamins, amino acids and salt. The only compounds that may escape destruction or denaturation are the salt solution, therefore you cant autoclave-sterilize your cell culture medium. You just need to filter it. This idea can be applied to any other solution you might want to sterilize in the future. You must be able to answer the question " How will this solution react to autoclave sterilizer temperature?". Whenever I want to sterilize, I make sure I give that question a befitting answer and I go ahead with the best option. You can do the same.
do you have to autoclave 30% sucrose in 1x pubs before you use it for cryoprotection for IHC?
I autoclave my solutions separately and there is no color change.Initially,i used to filter sterile my sugar solution with 0.2 micro filters but there was back pressure and sugar loss after filtration.As mentioned,after autoclaving, i added my phosphate and sugar solutions after cooling and my microorganisms just love it.
Is that mean, if i just autoclave the glucose solution, do i need to check the pH too?
Glucose is a reducing sugar and can react with amino acids in high temperatures (Maillard reaction).
The best way of doing it is to filter sterilize and add in the buffer/medium after autoclaving.
If you don't have sterile filters I would recommend to autoclave a glucose solution separately and mix it afterwards.
https://en.wikipedia.org/wiki/Maillard_reaction
You can autoclave it (separated) glucose 30%, and them mix with your autoclaved medium. I already did it (I am not sure if my glucose was 50%, but it was at least 30%). More than 50% for sure, it is not recommended. More than 50% could caramelize.
yes, autoclave glucose separately is good for microbiological media and work very well.
I prefer filter sterilisation to autoclaving for my glucose, the main goal is to get a sterile substrate which is quite possible with sterile 0.2 micro meter filters.
All the best for your experiments.
Glucose should not subjected to autoclave, because they very rapidly convert to 2 molecules of milk acid.
Only filter-sterlization.
you can add glucose and autoclave .otherwise u can add all other ingredients ,and autoclave the media ,and add filter sterilised glucose solution to this .
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