Science Buddies: "Ask an Expert"

Hello,

I am a current high schooler that plans on conducting research about the effect of ayurvedic medicine (specifically curcumin) on staphylococcus aureus (which causes eczema). Fortunately, I have a great deal of time before I can get started on this project. The problem is that I have no idea where to get started. Like I have absolutely no direction in which I'll be taking this project. I was hoping somebody had an idea on what I can do and where I can get started. If you are anyone who has any insight, please feel free to email me @[email protected]

Thank you so much

Hello,

This sounds like a great idea for a project! Microbiology is a really interesting field, and there are a lot of projects you can do with bacteria.

One of the first steps in planning this project is to decide what bacteria you want to use. Keep in mind that Staphylococcus aureus is a pathogenic organism, which means companies probably won’t sell or ship it to high schools or to individuals. You would only be able to use it if you worked on your project at a college or university. If you’re not able to do that, you can choose a different, non-pathogenic bacteria that’s safe for you to work with at your school or at home. I would recommend E. coli K-12. This is a safe E. coli strain that you can order from a science supply website like Home Science Tools or Carolina Biological. E. coli can cause skin infections in people who have eczema, so I think it’s still relevant to your project.

If this is your first time working with bacteria, you should familiarize yourself with the proper techniques and safety guidelines. Here are links to the Science Buddies Microorganisms Safety Guide and the Microbiology Techniques and Troubleshooting page that you can check out before you start your project.
https://www.sciencebuddies.org/science- ... sms-safety

https://www.sciencebuddies.org/science- ... leshooting

Now for the fun part: figuring out what experiments you want to do with the bacteria! One of the most common techniques for testing the effects of antimicrobial substances on bacteria is the Kirby-Bauer disk-diffusion method. This is a technique where you apply the antimicrobial substances to disks and then place the disks directly onto the surface of nutrient agar plates that have bacteria growing on them. If the bacteria die when exposed to the substance on the disk, then you know that it has antimicrobial properties. If you wanted to use this technique for your project, you could purchase liquid curcumin so that it will be absorbed by the disks. Here are the links to two Science Buddies projects that use the Kirby-Bauer method:

https://www.sciencebuddies.org/science- ... on#summary

https://www.sciencebuddies.org/science- ... on#summary

You could also browse the other Microbiology projects on Science Buddies to get some more inspiration:

https://www.sciencebuddies.org/science- ... crobiology

I hope this helps give you a place to start when planning your project. As you continue to plan, please feel free to post any other questions or concerns that you have!
Katelyn

Wow this was really insightful. Thank you so much! I also wanted to ask how exactly would I measure the decay rate of the bacteria. I'm having trouble thinking about what exactly should my dependent variable be. Should I stick with just curcumin or measure with a few other ayurvedic products such as neem powder, etc. What direction do you suggest I take this?

Once again, thank you!

Hello,

No problem, I'm glad it was helpful! If the antimicrobial substance on the disk is lethal to the bacteria, then the bacteria around the disk will start to die and form a zone of inhibition. This will look like a clear ring around the disk. If you measure the diameter of the zone of inhibition, this will tell you how susceptible the bacteria are to the antimicrobial substance. This procedure is described in more detail in the Procedure section of this project guide:

https://www.sciencebuddies.org/science- ... #procedure

I think it's a great idea to test out other ayurvedic products besides the curcumin! The filter disks are small so you could test out a few different products on the same agar plate. You can maybe choose 3 or 4 different products to test and see which one is the most effective as an antimicrobial.

I hope this helps!
Katelyn

Ok, thank you! This was all very helpful. I will let you know if anything else comes up when I actually start my experimentation!

Hey,

I had a quick question. Do you know where I can get Propionibacterium acnes from? I wanted to test the herbal medicine to see how they affect acne on the skin but I'm not sure how feasible it would be to acquire p acnes compared to e.coli k-12. Do you suggest I use P. acnes or E-coli?

Thank you!

Hi,

Propionibacterium acnes is going to be harder for you to obtain than E. coli K-12. It's available from ATCC, but you can't purchase it yourself as a student. You would need someone from your school to purchase it for you. You could talk to your teacher about whether or not that's a possibility.

https://www.atcc.org/Products/All/6919.aspx

However, Propionibacterium acnes is pretty difficult to culture- especially if you've never cultured any bacteria before. It needs to be grown in anaerobic conditions, so you would need an incubator with no oxygen inside. It also requires some special nutrients for growth. You can do some more background research on the exact method for culturing Propionibacterium acnes to see if it's something you could feasibly do. I think E. coli would be the better choice because it's much safer and easier to culture. It's a good option for students who are still learning how to culture bacteria, and it's easy to purchase it yourself from Home Science Tools or Carolina Biological.

I hope this helps!

Alright! But what would my driving factor be? Like does E-Coli also cause or possibly irritate acne and skin conditions?

Thanks for all the help!

Hello,

I don’t think E. coli plays a big role in acne specifically, but it does commonly cause other types of skin infections. One example is Gram-negative folliculitis. This is a disorder similar to acne that is caused by gram-negative bacteria like E. coli. E. coli can also infect skin wounds. This is a big concern for patients who are staying in the hospital after getting surgery. If you summarize what’s known about the role of E. coli in skin infections and why E. coli infection is a health problem that needs to be solved, I think that would provide a strong rationale for your project!

Something else you could do is do some research on the mechanism of action of each of your herbal medicines. How do they kill bacteria? Do they destroy the bacterial cell membrane? Do they inhibit some of the bacterial virulence factors? If the medicines target a structure that Propionibacterium acnes and E. coli both have, then you could reasonably argue that your results with E. coli would apply to Propionibacterium as well.

Are the medicines that you’re investigating for this experiment normally applied to the skin, or would they be consumed? E. coli is one of the most common bacteria found in the human intestines. If the herbal medicines are edible, this experiment could help you investigate what effect they might have on the gut microbiome. That’s just another avenue you could explore if you decide to move away from the skin.

I hope this gives you some inspiration! Let us know if you need more clarification on any of these points!

It's great that you said that because I was thinking of this the other day. E-coli actually causes UTI infections and since herbal medicines are consumable, I could find the effect of those herbal medicines on the human intestines as opposed to on the skin. Like turmeric is consumed when people have UTIs but I have not heard how effective it is. I think I am finally getting a good vision for this project. Thank you so much for everything!

And just to confirm, If I do the Kirky-Bauber test using the sterile disks, the herbs will need to be liquid, right?

That sounds like a great idea! I'm glad you were able to develop your vision for the project. Yes, the herbs would have to be liquid so that the disks can absorb them. The disks are made out of paper, so you just have to tap them on the surface of the liquid you want to test, and they will absorb it. If you have any more questions as you continue with your project, feel free to post them here!

Hello,

I'm finally gathering everything to start my project. I'm just having a little trouble finding liquid herbs. I know you can find them online but I wasn't exactly sure how pure the ones online would be. I read something about using centrifugation to separate water and the herbs to ensure that they are perfectly pure but I still don't think that the herbs that are left over as precipitates would be liquid. What should I do now? Should I just buy them or is there another method to finding liquid herbs?

Thank you so much for the help!

Hello again!

What specific herbs did you decide to use for your project? I can do some research online and see what's available. If you check the ingredients label on the liquid herbs and the only ingredients listed are the herb and the liquids it was dissolved in, I think it should be pure enough for your experiments.

I'm using Thyme, Curcumin, Ginger, and Holy Basil. Some of the online ones are pure and others are manipulated. That's why I thought it would just be a good idea to make them liquid myself. How should I do that?

Sure, I think it should be possible for you to prepare the liquid herbs yourself. You have to pay close attention to what the herbs are dissolved in because that could affect the results of your experiment. For example, curcumin is highly soluble in alcohols like ethanol, but ethanol by itself is antibacterial. That means that if you put herbs dissolved in ethanol onto your bacteria plates, you won't be able to tell if the bacteria are dying because of the alcohol or because of the herbs. For that reason, it would be best if you can get the herbs to dissolve in water. You can start by searching, "How to dissolve thyme in water," and repeat that for all your other herbs. If you pour hot water over the herbs and allow them to steep for a while, that should produce a liquid you can use for your experiment. Curcumin will probably be the most difficult since it's only partially soluble in water. If boiling water won't dissolve it, you can use dimethyl sulfoxide (DMSO). You should be able to buy it online.

Another option to consider would be to buy your herbs in essential oil form. Essential oils are usually extracted through steam distillation so they won't have alcohol in them and they will also be more concentrated than herbs dissolved in water. This might enhance their antimicrobial properties. They also should be pretty pure in this form. You could always try using both the herbs in water and essential oil herbs to see which ones kill the bacteria better.

Let me know if you have more questions!

Thank you so much! I've recently been looking into steam distillation. Do you think it would be a better idea to buy essential oils or make them yourself? It does look like a lengthy process but I also think making the essential oils myself shows more dedication from my end. Please let me know what you think.

You can definitely make them yourself if you want to! But keep in mind that distillation would require you to buy a still and those usually cost several hundred dollars. I think you can do steam distillation with a crock pot, but you would need enough of each herb to fill about half of the pot. If this doesn't seem do-able, I don't think there's anything wrong with just buying the essential oils. The main point of your project is studying the effects of the essential oils on bacteria, not making the oils themselves. So I don't think choosing to buy the oils would make you seem less dedicated! It's entirely up to you and what works the best with your schedule/budget.

Hello,

I have conducted my experiment numerous times and it fails every time. After about 8 trials, I am out of sterile discs, agar plates, and almost out of the bacteria. I have used an incubator and my E. Coli is growing fine. However, there is absolutely no zone of inhibition in any of my quadrants with the different herbal essential oils. I feel so discouraged because I have been planning this project for about 7 months now and the end result is that my project has failed. I don't have the time nor the money to buy new equipment and try the project again. I was supposed to write a research paper on this and submit it for a science fair but without any results, I don't know what I will even write it on. This project took a considerable amount of effort, especially because each essential oil was steam distilled in a high school chemistry lab, with each taking 6 hours and then cleaning the equipment after. It seems that effort has now failed. What should I do? Should I enter this failed project into the Science Fair?

Sorry for such a long message and thank you for everything!

Hello,

I'm sorry that the results of your project were disappointing! But your project definitely did not fail! All you did was get negative results. Your hypothesis was that your herbs would directly inhibit the growth of E. coli, but when you tested your hypothesis, you found that none of them inhibited E. coli growth. That's still super important information to know! You said yourself earlier that you heard of people using turmeric to treat UTIs, but you didn't know if it was effective or not. Now you know that, at least under your experimental conditions, it doesn't appear to be effective.

Negative results in science are extremely common. I work in a lab and I get a TON of negative results. I have also spent months working on expensive and time consuming experiments that "failed", so I totally understand your disappointment. A lot of people don't want to publish negative results because they think they're not interesting, but they're really so important. Reporting your negative results can save other researchers a ton of time and money on experiments that aren't going to give them the desired result.

You should absolutely enter your project into the science fair! The science fair is all about demonstrating your understanding of the scientific method and showing your enthusiasm for scientific research, and you clearly did both of those things! When you write your conclusion you can talk about why it was important to test your hypothesis and then you can go on to speculate about why you think the results didn't support your hypothesis. For example, you showed that the herbs don't directly inhibit E. coli growth, but maybe they could kill bacteria through a different mechanism. Perhaps they can activate the immune system when they're ingested and promote bacterial killing that way. You can also mention any technical issues that you think could have impacted the results or other alternative hypotheses you could test in the future. If you present your data clearly and logically and show that you followed the scientific method, your project will be viewed favorably! You showed a lot of amazing hard work, dedication, and critical thinking skills, and you should be proud of yourself!