UV-Vis Spectroscopy and Acid-Base Equilibrium in Bromothymol Blue

What is the goal of this experiment?

Determine the pKa of bromothymol blue using UV-Vis absorbance

What are the two main concepts used in this lab?

Acid-base equilibrium and UV-Vis spectroscopy

What color is the acidic form (HIn) of bromothymol blue?

Yellow

What color is the basic form (In⁻) of bromothymol blue?

Blue

Why does bromothymol blue change color?

Structural changes and electron delocalization when it gains or loses a proton

What happens when bromothymol blue loses a proton?

It becomes negatively charged (In⁻) and changes structure

What does a spectrophotometer measure?

Absorbance of light at different wavelengths

What is a spectrum?

A plot of absorbance vs. wavelength

What is λmax?

The wavelength where absorbance is highest

What is Beer's Law?

A = εbc

What does A represent in Beer's Law?

Absorbance

What does ε represent in Beer's Law?

Molar absorptivity (M⁻¹ cm⁻¹)

What does b represent in Beer's Law?

Path length (cm)

What does C represent in Beer's Law?

Concentration (M)

What is the key takeaway from Beer's Law?

Absorbance is directly proportional to concentration

What is Ka?

The acid dissociation constant

What does a larger Ka mean?

Stronger acid

What is pKa?

Negative log of Ka

What does a lower pKa indicate?

Stronger acid

What does a higher pKa indicate?

Weaker acid

What is the Henderson-Hasselbalch equation?

pH = pKa + log([A⁻]/[HA])

What happens at the inflection point?

[HIn] = [In⁻] and pH = pKa

What graph do you make first?

Absorbance vs. wavelength

What graph is used to determine pKa?

dAbsorbance/dpH vs. average pH

How do you find pKa from the derivative plot?

The peak corresponds to pKa

What is dAbsorbance/dpH?

The change in absorbance divided by change in pH

Why is the derivative used?

To locate the inflection point more precisely

What pH range is used in this experiment?

Approximately 5 to 10

Why are multiple pH values used?

To observe transition from acid to base form

What wavelength range is measured?

340-800 nm

At what wavelength are absorbance measurements taken in Part 2?

At the λmax of the basic form

What is used as the blank in spectrophotometer calibration?

DI water

Why is a blank used?

To set baseline absorbance to zero

What is a solvent-solvent extraction?

A separation based on differences in polarity or charge

Which layer is the aqueous layer?

The bottom layer

Which layer is the organic layer (ethyl acetate)?

The top layer

Which form of bromothymol blue is more soluble in water?

The charged basic form (In⁻)

Which form of bromothymol blue is more soluble in organic solvent?

The neutral acidic form (HIn)

Why are glass cuvettes used during extraction?

Ethyl acetate dissolves plastic cuvettes

Where should all waste be disposed?

In the liquid waste container (not the sink)

What happens to absorbance when concentration decreases?

Absorbance decreases proportionally

If a solution is diluted by 1/3, what happens to absorbance?

It becomes 1/3 of the original value

Which value indicates the strongest acid?

The largest Ka or smallest pKa

Which value indicates the weakest acid?

The smallest Ka or largest pKa

What does absorbance depend on?

Concentration, path length, and molar absorptivity

What happens when absorbance is very high (>2)?

Very little light reaches the detector

What is transmittance?

The fraction of light that passes through a sample

What is the relationship between absorbance and transmittance?

A = -log(T)

What is I₀ in absorbance equations?

Initial light intensity

What is I in absorbance equations?

Transmitted light intensity

What does the isosbestic point represent?

A wavelength where absorbance stays constant during conversion between forms

Why is bromothymol blue useful as an indicator?

Its acid and base forms have different colors

What happens at pH > pKa?

The basic form (In⁻) dominates

What happens at pH < pKa?

The acidic form (HIn) dominates

What type of acid is bromothymol blue?

A weak acid

What happens during acid dissociation?

HA ⇌ H₃O⁺ + A⁻

What determines solubility in extraction?

Polarity and charge

Which species prefers organic solvent?

Neutral/nonpolar species

Which species prefers aqueous solvent?

Charged/polar species

What is the purpose of the extraction in this lab?

To demonstrate solubility differences of acid/base forms

What type of data file is exported from the spectrophotometer?

.csv file

Why convert .csv to .xlsx?

To preserve graphs and formulas in Excel

What indicates the inflection point on a derivative graph?

The maximum peak