The Effects Of Drain Cleaner on Microbes Matt

The Effects Of Drain Cleaner on Microbes Matt Zagrocki Central Catholic High School Grade 9

Problem • What effect does drain cleaner have on the survivorship of Escherichia coli?

Purpose • This experiment was conducted to see the effect drain cleaner has on E. coli. • Will it decrease survivorship? • Does concentration have an effect? • Are eco friendly drain cleaners less damaging?

Application to the Real World • Drain cleaners are commonly used by the American household • Substances poured down drains can leak into the watershed • E. coli was used to model fresh water prokaryotes • Can a drain cleaner effectively sterilize household sinks and drains?

Drano Max Gel (industrial) • Widely used • Main ingredient is Sodium Hydroxide, a caustic • Other ingredients include sodium hypochlorite, (bleach) water, sodium silicate (corrosion inhibitor) and a surfactant blend (cleaning agent) • p. H of 11

Sodium Hydroxide • Molecular formula Na. OH • Highly reactive with other metals • Corrosive to living matter • Also referred to as lye • Synthetically manufactured

Green Gobbler (eco friendly) • Claimed to be biodegradable and eco-friendly • EPA approved • Unnamed hydroxide identified as “trade secret/proprietary” • Ph of 13 • Other ingredients are water, xanthan gum, alcohol ethoxylate, modified magnesium silicate

Escherichia coli • Common bacteria found in intestinal tract of animals • Works its way into water through feces • Gram Negative • Ubiquitous • classified as a facultative anaerobe • Thrives in p. H neutral environment • Survives in p. H range 4. 4 -9. 7

Hypothesis • Null Hypothesis – The drain cleaners will have no effect on E. Coli • Alternate Hypothesis – The drain cleaners will significantly reduce survivorship of E. coli • Green Gobbler will have less negative effects on E. coli survivorship than Drano

Materials • LB Agar Plates (0. 5% yeast extract, • Pipettes (p. 20, 200, and 1000) 1% tryptone, 1% sodium chloride) • %70 Ethanol (to sterilize) • E. coli • Permanent Marker • Drano Max Gel Drain Cleaner • Vortex • Green Gobbler Eco-Friendly Drain • Test Tubes (sterile) Cleaner • Test tube racks • Sterile Dilution Fluid (10 mm • Spreader Bars KH 2 PO 4, 1 mm Mg. SO 4, . 1 mm Ca. Cl 2, 100 mm Na. Cl)

Procedures 1. E. coli was grown overnight in sterile LB media. 2. Samples of the overnight cultures were added to fresh media in sterile sidearm flasks. 3. The cultures were placed in an incubator (37°C) until a density of 50 Klett spectrophotometer units was reached. This represents a cell density of approximately 108 cells/m. L. 4. The cultures were diluted in sterile dilution fluid to a concentration of approximately 105 cells/m. L.

Procedures (continued) 5. Six test tubes were set up as follows: 10% 1 1 1% 1 0. 01% 1 0. 1 Substock 0. 001% 0% 1 1 0. 01 0 Substock m. L of sterile dilution fluid 8 8. 99 9 Total volume 10 10 10 E. Coli m. L of drain cleaner

p. H values of Concentrations Stock 10% 1% . 001% Drano 11 ≈8. 1 ≈7 ≈7 Green Gobbler 13 ≈8. 3 ≈7 ≈7

Procedures (continued) 6. The tubes were vortexed and allowed to sit in room temperature for 10 minutes 7. 0. 1 m. L aliquots were pipetted onto agar plates 8. The plates were placed in an incubator for 24 hours, and then the colonies were counted visually. Each colony was assumed to have arisen from one cell.

E. Coli Survivorship in different concentrations of Drano 500 P =. 35 E-26 450 400 COLONIES 350 300 250 200 150 100 50 0 0% (control) 10^-3% Trial 1 Trial 2 0. 01% 0. 10% CONCENTRATION Trial 3 Trial 4 Trial 5 Trial 6 1% Average 10%

Dunnett’s Test- Drano P value < 0. 05 Concentration T Value Result 0. 001% 22. 23045 Significant 0. 01% 31. 1894 Significant 0. 1% 34. 86225307 Significant 10% 34. 86225307 Significant T crit – 3. 39

E. Coli Survivorship in different concentrations of Green Gobbler P = 4. 7 E-27 Green Gobbler 500 450 400 COLONIES 350 300 250 200 150 100 50 0 0% (control) 10^-3% Trial 1 0. 01% 0. 10% CONCENTRATION Trial 2 Trial 3 Trial 4 Trial 5 Trial 6 1% Average 10%

Dunnett’s Test – Green Gobbler P value < 0. 05 Concentration T Value Result 0. 001% 8. 4569076336 Significant 0. 01% 22. 2434517741 Significant 0. 1% 32. 459446345 Significant 1% 34. 86225307 Significant 10% 34. 86225307 Significant T crit – 3. 39

Average E. coli Survivorship in Different Concentrations of Drain Cleaner 500 P =. 35 E-26 450 P = 4. 7 E-27 400 COLONIES 350 300 250 P < 0. 05 200 150 P < 0. 05 100 P < 0. 05 50 0 0% (control) 10^-3% 0. 01% 0. 10% CONCENTRATION Drano Green Gobbler 1% 10%

Conclusions • Null Hypothesis was rejected • All concentrations with both drain cleaners had a significant effect • Drano appeared to have a greater negative impact at lower concentrations • Higher concentrations killed more colonies

Limitations • Only one bacteria was used • Only 5 concentrations were tested • Only two types of drain cleaner were tested • Plating, vortexing, pipetting, and exposure times could have been slightly unsynchronized • Study does not reveal other effects on bacterial health

Extensions • Use different types of bacteria • Use more types of drain cleaner • Use different exposure times • Additional trials per group

Sources • “DISSOLVE Drain Opener - Liquid Hair&Grease Clog Remover (32 oz. ). ” Green Gobbler, greengobbler. com/products/dissolve-liquid-hair-grease-clogremover? variant=12039958657. • “Max Gel Clog Remover. ” Drano® | SC Johnson, www. drano. com/enus/products/drano-max-gel-clog-remover. • “SODIUM HYDROXIDE. ” National Center for Biotechnology Information. Pub. Chem Compound Database, U. S. National Library of Medicine, pubchem. ncbi. nlm. nih. gov/compound/sodium_hydroxide. • “E. coli (Escherichia coli). ” Centers for Disease Control and Prevention, 25 Jan. 2018, www. cdc. gov/ecoli/index. html.

Anovas Anova- Single Factor / Drano SUMMARY Groups x. 1 x. 001 x. 0001 x 0 x (control) ANOVA Source of Variation Between Groups Within Groups Total Anova: Single Factor / Green Gobbler Count Sum 6 6 6 Average SUMMARY Groups x. 1 x. 001 x. 0001 x 0 x (control) Variance 0 0 0 0 0 274 45. 6666667 135. 466667 941 156. 833333 1920. 56667 2597 432. 833333 718. 566667 Count Sum 6 6 6 0 0 179 940 1967 2597 Average Variance 0 0 29. 8333333 156. 666667 327. 833333 432. 833333 0 0 190. 166667 773. 866667 1137. 36667 718. 566667 MS F ANOVA SS 880512. 556 13873 894385. 556 df MS F P-value F crit Source of Variation Between 5 176102. 511 380. 817079 3. 4886 E-26 2. 53355455 Groups Within Groups 30 462. 433333 35 Total SS 1024398. 47 df P-value F crit 5 204879. 694 435. 919396 4. 7456 E-27 2. 53355455 14099. 8333 30 469. 994444 1038498. 31 35