Science Buddies: "Ask an Expert"

Hi,

Controls are always good, and you should include a positive and a negative control in an experiment whenever possible, just to verify that your experimental protocol and reagents are all working as expected. For your experiment, the positive control could be the concentration of disinfectant that is recommended for use on the bottle label; the negative control would be the paper disc with no disinfectant. You would expect to see inhibition with the positive control and no inhibition with the negative control. Did you include these controls? If not, these please explain exactly what you did.

Don't forget to let me know the name of the disinfectant you are working with.

Donna Hardy

Wait, in your previous post, you say "I have a negative control with filter disks only dipped in distilled water. "

I thought you accidentally used tap water? I remember replying to one of your other topics, asking what you should do now that youve used tap water instead of distilled. Did you redo the experiment or something, using distilled water?

Sorry for the confusion.

The controls are dipped in distilled water.

However, when I made the concentrations I used tap water.

I realized the mistake shortly after making the concentrations.

Hi,

Tap water can have chlorine in it, which could potentially inhibit bacterial growth. However, your results showing 0 mm zone of inhibition suggests that this was not a problem. In the discussion section of your board, you will need to explain why the dilutions of disinfectant did not inhibit bacterial growth. One possibility is that the disinfectant is volatile and therefore evaporated before it could inhibit bacterial growth.

Did you have some more results to post?

Donna Hardy

Okay these are my final results.

Sorry, Donna I always seemed to forget to answer the questions about my disinfectants.
Details regarding that are included in the charts.

After this, I just need to type out what I did, my conclusion, discussion, and report.

Hopefully, the worst is over

Wait, you only had one Petri dish? I am asking because the website for your project on Sciencebuddies.com asks to use 20 petri dishes, 4 plates per disinfectant X 5 rounds of selection.

Remember that you are also trying to observe for signs of bacterial resistance--but that may prove to be rather difficult because you used a mixed source of bacteria, rather than one specific strain. Different types of bacteria become resistant to disinfectants at a different rate, some do not become resistant at all, some take hundreds of trials, etc.. It would be difficult to observe any bacterial resistance if you trimmed your # of rounds of selection to, say 2 or 3.

I know that sometimes, it takes tens and maybe even hundreds of times for certain bacteria to begin to show signs of bacterial resistance. I think that if you did not use at least 5 rounds of testing like the Sciencebuddies.com website did, you might not have a valid conclusion displaying bacterial resistance.

Edit: By the way, I looked online for a list of ingredients in the Clean & Clear astringent you used, and I found this: Active: Salicylic Acid (2%) Inactive: Alcohol, Benzophenone-4, Denatonium Benzoate, Dimethicone Propyl PG-Betaine, Fragrance, Isoceteth-20, Red 4, Water. I'm worried about the fact that there is alcohol present in the astringent. The presence of alcohol could have inhibited or promoted the development of the bacteria. I'm not sure about the other chemicals like Benzophenone-4, Denatonium Benzoate, Dimethicone Propyl PG-Betaine, but is it possible that you have time to quickly redo the experiment using an anti-acne product containing only salicylic acid and water?

I don't think so. The project is due January 7th, now. I don't have anymore petri dishes at my home either.

I'm thinking maybe I could just mention that in my discussion.

At this point, I'm just hoping for a good grade after all the hard work put in.

What do you think I should do now?

If you don't have enough time and/or enough petri dishes, then let it be. It's good that you're going to mention that in your discussion, but maybe you should ask your teacher first just in case he/she has a certain criteria they want specifically in each section.

Can you clarify the # of petri dishes you used for your experiments? Because like I mentioned before, the Sciencebuddies.com version of your project asks for 20 Petri dishes, 4 plates per disinfectant X 5 rounds of selection.

I used five disks.

The teacher misinformed me --telling me one round would be sufficient. I remember her saying that as long as I had a valid conclusion it would be fine.

Wait, so you used ONE Nutrient agar dish, but placed five paper disks on it? And that's it?

Wait, I'm so sorry. I made a mistake in my wording.

I used five petri dishes. Each petri dish had four sections.

I should probably go change my wording now on my results.

SORRY

Ok, I thought that's what you meant. I think you're done with the experiment part of your project now. You should focus on making the results organized and presentable. Remember, for your project board, to make the text large and readable from a distance. I think a minimum font size of 16 or 18 would be appropriate. Don't make your presentation board too colorful, because it looks as if you are trying too hard to impress. I think your project was quite advanced for your grade level, so do not worry that the results were not what you had expected.

Good luck

Okay. Your right.

You don't know how much I appreciate your help. I think if you hadn't of responded to my questions, I may have quit the project ages ago or right now had a panic attack.

Thanks.

Oh and one more thing.

What do you recommend out of the following for displaying my results.

Photos of petri dishes ( I managed to take clear quality photos on my cell phone camera rather than my digital --Strange, I know)

Or Pasting the Petri Dishes to the board..

I think you should paste them onto the board. Pictures, depending on the camera/lens you used, sometimes are not a clear representation of your petri dishes. Unless you have a top-notch, high-quality, professional photographer type camera, then use the pictures.

Oh, but couldn't the bacteria develop more from now to January 7th?

I was thinking of taking pictures..
and then additionally bringing in the disks.

I just want to prevent mishaps..i'm not sure if tape or any kind of glue is strong enough.

Good point, but even if the bacteria does continue to grow, it will not grow to a substantial amount that it will dramatically affect your results. The bacteria will not grow very much if you paste it on the board, because you are exposing it to temperatures which sway from cold to hot and hot to cold.

Use a very strong tape to tape the plates on the board. If you or your science lab has Parafilm, wrap the dish with Parafilm before pasting on the board.

Hi,

Jane has given you very good advice for presenting your project. Here are some additional comments that you might incorporate into your discussion section. This will help the science fair judges realize that you learned a lot in doing this project.

Thanks for posting the additional data and the identify of the disinfectant. It’s interesting that you obtained no inhibition of growth at all with even high concentrations of the disinfectant. The bacteria that grew on your plates were apparently not resistant at all to the product you used. The composition of the product is interesting. The alcohol would have inhibited the growth of bacteria initially, but is volatile so would have evaporated fairly quickly. The denatonium benzoate is a bitter compound that is added to denature alcohol, and the dimethicone propyl PG betaine and isoceteth-20 are surfactants so these would probably not significantly inhibit bacterial growth. And fragrance and red dye #4 have no known antimicrobial characteristics. The active ingredient in the product, salicylic acid, is described as a mild antiseptic, so it is possible that many bacteria are not inhibited by this compound.

http://en.wikipedia.org/wiki/Salicylic_acid

Salicylic acid works primarily by interfering with adhesion of bacteria to the surfaces. This mechanism could help prevent a clinical infection, but would not necessarily kill the bacteria.

http://www.jstor.org/pss/30107422

It would be interesting to know how the manufacturer evaluated the effectiveness of the product, and you may want to look at the product claims on the label and then ask the manufacturer how the antimicrobial activity was measured.

You worked with unknown bacteria. It would be interesting to repeat this experiment with Propionibacterium acnes to compare results. Or, if you aren’t able to do any more lab work, at least suggest this possibility in your discussion.

Donna Hardy

Parafilm....Hm.. I'm not sure what it is.

But I'll look into it.

I'll ask my teacher on Monday.

I came to the conclusion that "less is more." That in the sense the more diluted the disinfectant solution was the more bacteria was killed.

How do you think I can support this scientifically and with research?

Do you have any helpful links or ways that I can show that this is possible?

Thanks. I tried to research it, but I didn't come up with much. Do you know how I can go about this?

Both the Lucky Soft Soap and Acne product had messed up results and contained Benzophenone 4. Is it safe to conclude that this could have caused an issue? Thanks guys!

this is my conclusion:

My hypothesis was that the dilution of 12.5% of the disinfectants would inhibit or prevent the most bacterial growth compared to the dilutions of 25% and 50%. My results partially support my hypothesis. For the products of mouthwash and the all purpose household cleaner my inference was true. When mouthwash had a concentration of 50% there were two bacterial colonies, however for the concentration of 12.5% there was only one. Additionally, for the all purpose household cleaner there were four colonies at the dilution of 50%, but only one in the section for the dilution of 12.5%. This probably occurred because the lower concentrations are more effective in disrupting the cell membrane. However, that was not the case for the soap and anti-acne product – for them, most bacteria was destroyed at the level of 25%. Also, at the same time, though, the anti-acne product contained alcohol which can either inhibit or promote bacterial growth; thus, distorting my results. Furthermore, both the Lucky Soft Soap and Clean & Clear anti-acne product contain benzophenone -4, leading me to believe that this could have possibly altered my results. Nonetheless, based on my research and experimental results and findings I can conclude that the less concentrated a solution of disinfectant is the more bacterial growth it will inhibit.

I'm not too sure about talking about the alcohol in the anti-acne product, because the mouthwash contained it too.

I'm trying to think of what substance could have distorted my results.

Hi,

I am not exactly sure if Benzophenone-4 may have affected your results--I'll leave that part to the experts. I just wanted to say, your hypothesis seems very long. I've been doing science fair projects since elementary school, and the conclusion is generally the part where you only say My hypothesis that ___ was partially incorrect. For the products of mouthwash and the all purpose household cleaner my inference was true. When mouthwash had a concentration of 50% there were two bacterial colonies, however for the concentration of 12.5% there was only one. Additionally, for the all purpose household cleaner there were four colonies at the dilution of 50%, but only one in the section for the dilution of 12.5%. The rest of your conclusion you posted should be omitted, or if you have space, add it to the discussion.

Remember you don't want to have just an array of text--those projects are very unappealing. For the conclusion, the portion I italicized above should be sufficient. For the discussion, I know that school science fair projects should have just half a page of text, maybe a tad bit more. On the board, just lightly touch on the things that you wish to mention in the report. You don't want to bore the reader with words..

(Btw, sorry if I went a little off topic )

Oh Okay. I understand what your saying. However, the teacher said "1 paragraph." All the years before, I always wrote two sentences.
I'll try to shorten my conclusion and maybe bring it to the point faster. I used the science buddies format for a conclusion.

Once I fix my conclusion, I'll put it up again.

I revised my conclusion.

My hypothesis that the dilution of 12.5% of the disinfectants would inhibit or prevent the most bacterial growth compared to the dilutions of 25% and 50% was partially correct. For the products of mouthwash and the all purpose household cleaner my inference was true. When mouthwash had a concentration of 50% there were two bacterial colonies, however for the concentration of 12.5% there was only one. Additionally, for the all purpose household cleaner there were four colonies at the dilution of 50%, but only one in the section for the dilution of 12.5%. This probably occurred because the lower concentrations are more effective in disrupting the cell membrane. However, that was not the case for the soap and anti-acne product – for them, most bacteria was destroyed at the level of 25%. Nonetheless, based on my research and experimental results and findings I can conclude that the less concentrated a solution of disinfectant is the more bacterial growth it will inhibit.

Better? Worse? Comments?

And this is my abstract, if it helps your understanding more. I realized mid-experiment that my experiment more tested concentration levels rather than bacterial resistance development over time.
The teacher advised 1 to 2 paragraphs. I felt one paragraph was enough, because I didn't want to bore the reader.

Advertisers of disinfectants are always touting new cleaning products which are marketed as highly concentrated and claim to enhance your results emphatically --but really what good are they serving you when they fill your home with harmful amounts of powerful substances? In my experiment I tested altered dilutions of four types of disinfectants (antibacterial soap, all purpose household cleaner, mouthwash, anti-acne product) to apprehend what concentration level will eminently inhibit bacterial growth, or in other words which concentration level is most effective. By conducting this project I hoped to determine that you can use dilute the amount of disinfectant utilized and still be able to have the most adequate results. In order to test my hypothesis, I set up petri dishes employing the disk diffusion method in which I tested the positive control (100% of a disinfectant), a negative control (distilled water), and dilutions of 50%, 25%, and 12.5% of disinfectants (independent variable). The disinfectant would diffuse into the medium and thus killing some portion of the common bacteria (swabbed from a door knob in my home) inoculated on the surface of the agar. After I calculated how many bacterial colonies (dependent variable) survived at each concentration level in order to come to my conclusion, I established that my hypothesis was did considerably reflect the outcome to my experiment with a few exceptions.

You should omit in your conclusion, or say it during your presentation: This probably occurred because the lower concentrations are more effective in disrupting the cell membrane. However, that was not the case for the soap and anti-acne product – for them, most bacteria was destroyed at the level of 25%.

I realize that the Sciencebuddies website combines Conclusion & Discussion into one large category. I remember, as a teenager, always separating the Conclusion and Discussion sections. I don't know if thats how your school does it, but if so, you should leave out this part from the Sciencebuddies conclusion criteria:

If appropriate, state the relationship between the independent and dependent variable.

Summarize and evaluate your experimental procedure, making comments about its success and effectiveness.

Suggest changes in the experimental procedure (or design) and/or possibilities for further study.

For your abstract, you are not using the disk diffusion method. The disk diffusion method only works when you are testing ONE strain of bacteria. You used a mixed culture from your doorknob. Also, the disk diffusion method tests the zone of inhibition, and your experiment tested the # of bacteria colonies. So you should fix the part where you say you used the disk diffusion method--maybe just say you simply counted the # of colonies.

By the way, I don't think you should use large/sophisticated vocabulary words, especially in the eighth grade. Even if you do know what those words mean w/o a thesaurus, it will bore your peers when you read it. Trust me; I made the same mistake. Like your project proves, less is more.

Hi,

Thanks for posting your abstract. I have looked at your data again, and I don't think it is possible to make conclusions you are proposing, and here are the reasons:

1. The number of colonies in your controls was 12, 3, and 13. The number of colonies in your test sections ranged from 1 to 23 colonies. Generally, microbiologists don't count Petri dishes with less than 30 colonies, so it is not possible to say if there is any statistically significant difference in the number of bacteria between the controls and test sections.

2. The method of measuring the effect of antimicrobial agents you are using, by measuring the distance between the edge of a filter paper disk that has been soaked in the antimicrobial agent and the closest colonies on the plate. All of your zones of inhibition, with the exception of the bleach were 0 mm. That means that the bacteria were able to grow up to edge of the paper disk and were not inhibited at all. There is essentially no difference between any of your results. I don't know if the 4 mm zone of inhibition you observed with the bleach is significant.

3. The zones of inhibition test is normally done using about 10,000 bacteria/plate of a pure culture of bacteria that are in log phase growth. You used a mixed culture source of unknown bacteria in an unknown phase of growth, so inadvertently added a lot of variables to your experiment. Your experiment would have been better controlled if you had used a single bacterium.

4. Normally, the zones of inhibition are measured at 24 hours of growth. You did not observe any growth for a few days, and this may have allowed the chemicals in the paper disks to deteriorate. I'm sure the alcohol in the mouthwash and acne product evaporated; the bleach may have been reduced.

5. The salicylic acid in the anti-acne product does not kill bacteria, but inhibits them from sticking to surfaces or making biofilms. This anti-microbial effect cannot be measured by testing for zones of inhibition. There are assays for measuring biofilms, and it would be possible to measure anti-biofilm activity by using a different type of assay.

6. You conducted your experiment one time and included one test disk per product (if I am interpreting your results correctly). It would have been better to test each product in duplicate and repeated the entire experiment.

I recommend at the beginning of your conclusion that your hypothesis was not confirmed or disapproved, because your results are inconclusive. You can then proceed with an explanation of what happened and what you would do if you had time and materials to repeat the experiment.

Donna Hardy