vanillabean16
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testing for flavonoids in tea with milk. oh dear, help.

Postby vanillabean16 » Sun Mar 11, 2007 3:27 pm

hello anybody and everybody.

i am a junior in high school, and i'm doing the science fair. i'd really love to somehow test the theory that adding milk to green tea takes away the healthful benefits. I have found many articles saying that adding milk, which has caseins, to tea, which has catechins, decreases the concentration of catechins, thus taking away its anti-cancer benefits.

So. i basically need a way to test for levels of catechins in a liquid, so i can prove that adding milk decreases their concentrations in tea. i don't know if its at all possible, but it would be incredible if anybody could give me some help!

thank you thank you :)

Louise
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Re: testing for flavonoids in tea with milk. oh dear, help.

Postby Louise » Sun Mar 11, 2007 4:54 pm

vanillabean16 wrote:hello anybody and everybody.

i am a junior in high school, and i'm doing the science fair. i'd really love to somehow test the theory that adding milk to green tea takes away the healthful benefits. I have found many articles saying that adding milk, which has caseins, to tea, which has catechins, decreases the concentration of catechins, thus taking away its anti-cancer benefits.

So. i basically need a way to test for levels of catechins in a liquid, so i can prove that adding milk decreases their concentrations in tea. i don't know if its at all possible, but it would be incredible if anybody could give me some help!

thank you thank you :)


Have you tried googling for catechin assay or catechin testing? I found tons of papers that listed procedures. Many used an instrument called an HPLC, which I am guessing you don't have access too, but there may be some chemical assays as well.

Catechin levels in green tea are widely measured, so I am sure you will find something that will work with what you have.

I have found many articles saying that adding milk, which has caseins, to tea, which has catechins, decreases the concentration of catechins, thus taking away its anti-cancer benefits.


And how did these papers do the testing?

If you want to ask questions about a specific procedure or ask for us to explain the science lingo after you've done some research, feel free to post back

Louise

vanillabean16
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Joined: Sun Mar 11, 2007 3:18 pm

Postby vanillabean16 » Mon Mar 12, 2007 1:51 pm

thanks for writing back!

i looked at a bunch of procedures for catechin testing, and they ALL involved using HPLCs, and you're right, i don't have access to one. However, i may have access to a spectrophotometer. I have seen this option mentioned many places, but i can't find any procedures that actually describe how this can be done.
Do you know of any?

Thank you SO much!!
-Emily.

Louise
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Postby Louise » Mon Mar 12, 2007 2:47 pm

vanillabean16 wrote:thanks for writing back!

i looked at a bunch of procedures for catechin testing, and they ALL involved using HPLCs, and you're right, i don't have access to one. However, i may have access to a spectrophotometer. I have seen this option mentioned many places, but i can't find any procedures that actually describe how this can be done.
Do you know of any?

Thank you SO much!!
-Emily.


I think it is used with an HPLC. HPLC stands for high pressure (performance?) liquid chromatography, and it is the way you separate out the different components. It is like a tube filled with little beads. Some chemicals stick to the beads and others don't. So, some chemicals go through very quickly (because they don't stick) and others very slowly (because they do stick). So, at each time, you have a different chemical coming out of the tube. You then use a spectrophotometer to tell what the identity of each chemical is.

I will look around a little more, but I was thinking about this last night, and you may be out of luck. HPLC is one of the most widely used modern analytical instruments, and most of the green tea research is pretty recent, so it may be that all the papers use HPLC.


Louise

Louise
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Postby Louise » Mon Mar 12, 2007 4:01 pm

Louise wrote:
vanillabean16 wrote:thanks for writing back!

i looked at a bunch of procedures for catechin testing, and they ALL involved using HPLCs, and you're right, i don't have access to one. However, i may have access to a spectrophotometer. I have seen this option mentioned many places, but i can't find any procedures that actually describe how this can be done.
Do you know of any?

Thank you SO much!!
-Emily.


I think it is used with an HPLC. HPLC stands for high pressure (performance?) liquid chromatography, and it is the way you separate out the different components. It is like a tube filled with little beads. Some chemicals stick to the beads and others don't. So, some chemicals go through very quickly (because they don't stick) and others very slowly (because they do stick). So, at each time, you have a different chemical coming out of the tube. You then use a spectrophotometer to tell what the identity of each chemical is.

I will look around a little more, but I was thinking about this last night, and you may be out of luck. HPLC is one of the most widely used modern analytical instruments, and most of the green tea research is pretty recent, so it may be that all the papers use HPLC.


Louise


I found a reference that might be useful;
From:http://www.worldnutra.com/Newsletter_Papers/Quantification_and_Evaluation_of_Antioxidants.html

"phenolic compounds such as catechins and the components of Echinacea spp. have been found to have excellent antioxidant properties and are linked to a wide range of health benefits. To determine total polyphenolic levels a spectrophotometric method is employed(1). In this assay, samples are extracted with methanol and Folin-Ciocalteu reagent is added. The samples are then placed in a spectrophotometer and the absorbance is compared to a gallic acid reference standard. Results are presented as gallic acid equivalents (GAE)."
[1] Sakakibara, H., Honda, Y., Nakagawa, S., Ashida, H., and Kanazawa, K. "Simultaneous Determination of all Polyphenols in Vegetables, Fruits, and Teas," J. Agric. Food Chem. 51, 572-580 (2003).

This will tell you about total polyphenol levels. I have no idea what these chemicals are. I wil look at the original paper at the university tomorrow.

Louise

vanillabean16
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Postby vanillabean16 » Mon Mar 12, 2007 5:14 pm

thanks, that did help. i guess the fact that high performance liquid chromatography is used in conjunction with spectrophotometry accounts for my inability to find any procedures!
i suppose my next step is to find the simplest procedure possible, and to somehow get my hands on those two instruments. Perhaps they could be available for rent commercially, or by local universities?
i wonder why i just HAD to pick the most complicated experiment possible.....

Emily.

vanillabean16
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Joined: Sun Mar 11, 2007 3:18 pm

Postby vanillabean16 » Mon Mar 12, 2007 5:24 pm

i just found this:

http://pubs.acs.org/cgi-bin/abstract.cg ... 0855a.html

entitled: Interactions between Flavonoids and Proteins: effect on the total antioxidant capacity.

it uses a spectrophotometer, and some confusing chemicals.
But this is closer than anything else i have found.
what do you think?

Louise
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Postby Louise » Mon Mar 12, 2007 5:30 pm

vanillabean16 wrote:thanks, that did help. i guess the fact that high performance liquid chromatography is used in conjunction with spectrophotometry accounts for my inability to find any procedures!
i suppose my next step is to find the simplest procedure possible, and to somehow get my hands on those two instruments. Perhaps they could be available for rent commercially, or by local universities?
i wonder why i just HAD to pick the most complicated experiment possible.....

Emily.


This last article I posted did not use HPLC, but it only measures total polyphenols with a spectrophotometer and two chemicals as indicators and standards.

The HPLC is fairly expensive (I'm guessing $100,000 for a decent system) and is very complicated to use. While the local university probably has many, they probably won't share. :) A spectrophotometer is also expensive (but much cheaper than an HPLC) and MUCH easier to use (and much harder to screw up if you don't know what you are doing.

These instruments are probably not rentable, and for the HPLC at least, you couldn't operate it even if you had one.

I will try to get the procedure from the Journal of Agricultural Food Chemistry tomorrow. I think that is your best bet... or a new project. :(

Louise

Louise
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Postby Louise » Mon Mar 12, 2007 5:50 pm

Louise wrote:
vanillabean16 wrote:thanks, that did help. i guess the fact that high performance liquid chromatography is used in conjunction with spectrophotometry accounts for my inability to find any procedures!
i suppose my next step is to find the simplest procedure possible, and to somehow get my hands on those two instruments. Perhaps they could be available for rent commercially, or by local universities?
i wonder why i just HAD to pick the most complicated experiment possible.....

Emily.


This last article I posted did not use HPLC, but it only measures total polyphenols with a spectrophotometer and two chemicals as indicators and standards.

The HPLC is fairly expensive (I'm guessing $100,000 for a decent system) and is very complicated to use. While the local university probably has many, they probably won't share. :) A spectrophotometer is also expensive (but much cheaper than an HPLC) and MUCH easier to use (and much harder to screw up if you don't know what you are doing.

These instruments are probably not rentable, and for the HPLC at least, you couldn't operate it even if you had one.

I will try to get the procedure from the Journal of Agricultural Food Chemistry tomorrow. I think that is your best bet... or a new project. :(

Louise


Looks like "Journal of Agricultural Food Chemistry" likes tea. I posted a citation to this journal (different article) a couple of posts ago. The one you linked to is confusing. Maybe the one I found will be clearer.

would your teacher be able to order chemicals for you?

Louise

vanillabean16
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Postby vanillabean16 » Mon Mar 12, 2007 5:55 pm

i am going to ask her tomorrow if she can order the chemicals, and if the procedure is do-able. Plus, my father is the head of the ER at one of the local hospitals, so he may be able to find connections through the pharmacy. He also thinks that the hospital might have a spectrophotometer. My school has one, but it is from the early 80s, and nobody knows how to work it! But i'm working on trying to find directions through its barely visible serial number :)

Craig_Bridge
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Postby Craig_Bridge » Mon Mar 12, 2007 5:55 pm

HPLC and Spectography equipment is not something you can rent. Long term lease maybe, but not very affordable.

The care of this equipment and the columns isn't something you just let anybody do. My wife baby sat this kind of equipment for a pharmacy professor for a couple of years as an undergrad lab assistant.

See if somebody at a local university does similar research and will help you out. If not, then probably need a different project or methodology.
-Craig

Louise
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Postby Louise » Mon Mar 12, 2007 6:02 pm

vanillabean16 wrote:i am going to ask her tomorrow if she can order the chemicals, and if the procedure is do-able. Plus, my father is the head of the ER at one of the local hospitals, so he may be able to find connections through the pharmacy. He also thinks that the hospital might have a spectrophotometer. My school has one, but it is from the early 80s, and nobody knows how to work it! But i'm working on trying to find directions through its barely visible serial number :)


If you can tell me the name and the brand and the model number of the spectrophotometer at school, I might be able to tell you how to use it.

If this is a single wavelength mesurement, then we might be able to construct an instrument with filter glass and a photodiode.

Louise

vanillabean16
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Postby vanillabean16 » Mon Mar 12, 2007 7:11 pm

the spectrophotometer is a Bausch & Lomb Spectronic 20. The only number readable on the back said: CAT NQ. 33-29-95
I typed just its model name and type into google and found it easily. Its pretty old! which means i bet its a sinch to use. that is, if its still working...

Louise
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Postby Louise » Mon Mar 12, 2007 7:37 pm

vanillabean16 wrote:the spectrophotometer is a Bausch & Lomb Spectronic 20. The only number readable on the back said: CAT NQ. 33-29-95
I typed just its model name and type into google and found it easily. Its pretty old! which means i bet its a sinch to use. that is, if its still working...


Ah, a Spec 20. Those things are pretty durable. This is the "analog" not "digital".

I'm guessing you found a bunch of instructions on the web- they are pretty simple. The first thing to do is see if the lamp works.

You want to turn it on- following the appropriate warmup of 15 minutes, etc. Set the wavelength to 532 nm. Look in the sample comparment. Carefully stick a white piece of paper in the sample cell. You should see a green light. Change to 640 nm, should see red light. If this checks out, you are good to go except for the chemicals, I think. If you see the wrong color light, just write down what color, and we can try to calibrate it.

If no light, then we have a problem... ask if they have an extra tungsten lamp.

Louise

tdaly
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Postby tdaly » Mon Mar 12, 2007 9:51 pm

What part of the country do you live in? An organization that I work with has a lab dedicated to high school science fairs, and we have all sorts of things: infared spectrophotomers, UV-VIS spectrophotometers, HPLC, GC-MS, etc. We also have the ability to handle most "nasty" chemicals and materials. Let me know the general area where you live, and I can tell you if using this lab might be a possibility.
All the best,
Terik


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