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The relationship between acid pH and pKa (acid dissociation constant) in aqueous solutions is described by the Henderson-Hasselbalch equation. This equation can be used to forecast the pH of a buffer solution when the concentrations of such acid as well as its conjugate base, or the base and the related conjugate acid, are determined.

Henderson - Hasselbalch equation relates pH with p K a$\mathrm{as}$

$\mathrm{pH}=\mathrm{pKa}+{\log }_{10}\left(\frac{\left[\mathrm{A}\right]}{\left[\mathrm{HA}\right]}\right)$

The pH of the microscopic vesicles is determined by infusing an indicator into the living cells. The indicator quotient$\frac{\left[{\mathrm{I}}^{-}\right]}{\left[\mathrm{HIn}\right]}$is measured spectroscopically. Using the measurement of indicator quotient obtained by spectroscopy, the Henderson – Hassel Balch equation for this case is written and pH is calculated as,

role="math" localid="1663399695447" $\mathrm{pH}=\mathrm{pKa}+{\log }_{10}\left(\frac{\left[\mathrm{In}\right]}{\left[\mathrm{HIn}\right]}\right)$

The usage of spectrochemical data in determining the pH of microscopic vesicles was explained.