The Effect of Mediterranean Aqueous Herbal Extracts on Escherichia coli K12 Population Growth

Abstract:

Over time many cultures have used a variety of herbs and spices as ways of not only flavoring foods, but also as a bacteriostatic to reduce bacterial growth and prevent spoilage.  Herbs and spices are often combined together into mixes, like curries, za’atar, and Mediterranean blends, suggesting that herbs and spices may work synergistically.  Aqueous extracts of basil, oregano, rosemary, and thyme were made as follows.  1g of dried material was ground using a mortar and pestle and then diluted in 25 ml of sterile water.  The solution was set aside for 24 hours and then centrifuged at 4000 xg for 10 minutes.  The supernatant was removed and then passed through vacuum filtration.  This fluid was then sterile filtered using a 0.2 µm filter.  Sterile aqueous extracts were combined with equal parts to produce all combinations of one, two, three, and four spices.  Overnight cultures of Escherichia coli K12 were diluted in tryptic soy broth (TSB) to produce cultures of 1 x 10^8 cells/ml.  In the first experiment, a 96-well plate was used to establish E. coli populations consisting of 1% spice extracts in all combinations, while in the second experiment, 10% spice extracts were used.  Both experiments also consisted of wells that acted as blanks.  Well absorbances were measured using a plate reader set to measure the absorbance at 600 nm every 30 minutes for 36 total hours.  Methodologies were explored to calculate r (intrinsic rate of increase) and K (carrying capacity) from the logistic growth equation.  The R package, GrowthCurver, was also used to calculate r and K, as well as another measure, area under the curve.  We found that in the 10% extracts, oregano consistently reduced K and area under the curve.