Heartburn

Example 5

The content of free fatty acids in the oils and products was determined by neutralization titrimetry. The free fatty acids, about 0.2 g of the sample, were titrated with 0.04 mol·L⁻¹ NaOH solution in a Mettler model DG20 automatic titrator up to a pH of 11.0 and the acidity of the sample was determined from Equation 3.

Alternatively, for samples with a larger volume, the free fatty acids, from about 0.5 to 1 g of the sample, were titrated with 0.25 mol·L⁻¹ NaOH solution using phenolphthalein as indicator and the acidity of the sample was determined from Equation 3.

$\begin{array}{cc}\mathrm{Acidity}\left(\%w\text{/}w\right)=\frac{V\times M\times \mathrm{AG}}{10\times m}& \left(\mathrm{Equation}3\right)\end{array}$where:

• • V=volume of sodium hydroxide used in titration of the sample (mL);
  • M=molarity of the NaOH solution (mol·L⁻¹);
  • AG=molecular weight of the fatty acid present in highest concentration in the oil* (g);
  • m=sample weight (g).
  • *Soya oil=linoleic acid (280 g); castor oil=ricinoleic acid (298 g).

Example 5

The Lactobacillus ingested through the oral cavity passes through the stomach with the lower acidity and the intestines with high digestive enzymes and are exposed to low pH of gastric acid, pepsin, intestinal bile salts and digestive enzymes. Therefore, in order to utilize microorganisms as probiotics, gastric juice resistance is essential to survive in low pH and enzymes, and bile juice resistance is essential to survive in extreme intestinal environment. In accordance with the present disclosure, experiments were conducted to identify resistance to artificial gastric juice and bile juice of the above two strains with superior inhibitory effects against Gardnerella vaginalis and Candida albicans. The pH of the gastric juice in the body is maintained at about 3.0, and the food passes through the stomach for about 3 hours. In general, when maintaining viable cell count for 3 hours or more at pH 3, the cells has the high resistance to acidity. In order to identify the intestinal viability of Lactobacillus, survival experiments for artificial gastric juice and artificial bile juice were conducted with reference to Maragkoudakis' method. MG4272 and MG4288 strains were streaked on MRS plate medium and incubated at 37° C. for 24 hours, and the resulting colonies were inoculated in MRS liquid medium and incubated (37° C., 24 hours). Then, 2% passage was incubated for 24 hours in fresh MRS medium. The culture medium was then centrifuged (4,000×g, 4° C., 5 minutes) and washed twice with phosphate-buffer saline (PBS, pH 7.4). The washed cells were adjusted to OD₆₀₀ 1.0 (10⁸ to 10⁹ CFU/mL) and used for resistance experiments to the artificial gastric juice and artificial bile solution, respectively. As a control, 900 μL of pH 7 PBS was added to 100 μL of diluted Lactobacillus and the mixture was shaken and the number of viable cells was measured immediately. In order to identify the resistance to gastric juice, pepsin (Sigma-Aldrich, Saint Louise, USA) was dissolved in 3 g/L of pH 3 to pH 4 PBS to prepare an artificial gastric juice. 100 μL of lactobacillus diluent was added to 900 μL of artificial gastric juice, shaken, and cultured at 37° C. In 3 hours, the viable cell count was measured. To identify resistance to the artificial bile juice, pancreatin (Sigma-Aldrich, Saint Louise, USA) was dissolved in 1 g/L at pH 7 to pH 8 to prepare artificial bile juice. 100 μL of lactobacillus diluent was added to 900 μL of artificial bile juice, shaken and incubated at 37° C. In 4 hours, the viable cell count was measured. The measured results are shown in Table 1 in terms of log CFU/ml.

As shown in Table 1 both strains of MG4272 and MG4288 were identified to maintain the viable cell count of 10⁸ CFU/mL or more after 3 hours at pH 3, thereby identifying excellent acid resistance. In the artificial bile resistance test, both strains of MG4272 and MG4288 were identified to maintain the viable cell count of 10⁸ CFU/mL or more, thereby identifying excellent bile resistance.

Example 7

The conversion of free fatty acids to biolubricants was monitored by the titrimetric method (acidity), in which the consumption of the FFAs was observed. The conversion of the reaction was calculated from Equation 5 or Equation 6.

The results from Equation 5 are expressed in the form of percentage of FFA esterified disregarding the excess of FFA in the reactions, so that 100% conversion is equivalent to 100% of the hydroxyls of the alcohol being esterified. The results from Equation 6 are expressed in the form of percentage of FFA esterified, so that 100% conversion is equivalent to 100% of the FFAs esterified.

$\begin{array}{cc}\mathrm{Conversion}\left(\%\mathrm{hydroxyls}\right)=\frac{100\times \left(A_i-A_f\right)\times \mathrm{RM}}{H\times A_i}& \left(\mathrm{Equation}5\right)\\ \mathrm{Conversion}\left(\%\mathrm{FFA}\right)=\frac{100\times \left(A_i-A_f\right)}{A_i}& \left(\mathrm{Equation}6\right)\end{array}$where:

• • A_i=initial acidity (% w/w);
  • A_f=final acidity (% w/w);
  • RM=initial molar ratio FFA/alcohol;
  • H=number of hydroxyls in the alcohol molecule