Ch 8. Spectrophotometry Flashcards
Does photodiodes can give us the exact wavelength of the light or the exact color or color combination of a light signal? why?
No, it is just sensible to a wavelength range.
what technique/s is used for get detailed information of a light signal?
Spectophotometry
How current spectrophotometer are different from the old versions of the device?
old ones. bulky and expensive. Not considered for biosensors
current ones- small, and cheaper. becoming an integral part
of many sensor and biosensor devices.
what does spectrophotometry measure?
The light intensity absorbed or transmitted through a sample material. (liquid or gas)
what is photometry?
the intensity measurement of a specific wavelength.
what is spectrometry?
The measured intensity through a range of wavelengths.
What is the primary function of a spectrophotometer?
They typically measure the light intensity absorbed , transmitted, scattered, or reflected by a sample at specific wavelengths. generating an absorption spectrum
what are UV/Vis spectrophotometer?
The most common type of spectrophotometer uses UV and Vis light.
what is a FT-IR spectrophotometer?
Measures the absorption of IR light
how does light absorbance (A) is equated?
A=-log I/I_0
how is transmittance (T) equated?
T=I/I_0 | A=-log(T)
Why Absorbance is more common than transmittance?
it is related to the concentration of a specific component. A=εlc
ε :molar absorptivity
l : path length (the distance the light travels through the material)
c : molar concentration of a specific component
Beer-Lambert law.
if we have a solid sample, can be measure its absorbance?
No. In this case, only can be measured the intensity of reflected light using a reflectometer.
What is the difference between and spectrophotometer and a reflectometer?
The main difference is the change in the photodiode placement. A photodiode in a reflectometer is on the same side as a light source. In contrast, a spectrophotometer has its photodiode placed in front of the light source.
what is a absorption spectrum?
it is a plot of A-λ. A is recorded continuously while varying λ. A graph showing how much light a sample absorbs at different wavelengths.
what are the parameter that an oximeter measures?
Oxygen saturation in blood (SpO2%) and heart rate (pulse)
What is spectrophotometry?
A technique that measures the intensity of light absorbed or transmitted through a sample material, often as a function of wavelength
What physiological parameters can a pulse oximeter measure?
Pulse oximeters measure blood oxygen saturation (SpO2) and pulse rate
Define SpO2.
SpO2 is the percentage of oxygen-saturated hemoglobin relative to the total hemoglobin in the blood
Sketch the absorbance spectra of oxyhemoglobin and deoxyhemoglobin. Identify two wavelengths used by a pulse oximeter.
Oxyhemoglobin absorbs more at ~940 nm (infrared), while deoxyhemoglobin absorbs more at ~660 nm (red). These wavelengths are used for SpO2 calculation
What wavelengths does a pulse oximeter typically use?
Red (~660 nm) and infrared (~940 nm).
Describe the principle behind hemoglobin quantification using spectrophotometry.
Different forms of hemoglobin (oxy-, deoxy-, met-) have unique absorbance peaks that can be used for identification and quantification.
What is numerical aperture (NA) in optical fibers?
NA is the sine of the half-angle of the light acceptance cone, defining the fiber’s light-gathering ability.
Why is a miniature spectrophotometer advantageous?
Compact size, affordability, and versatility make it suitable for portable and field applications.
How does light scattering affect spectrophotometric measurements?
It can distort absorbance readings, necessitating techniques like 90° or 180° fluorescence detection to minimize interference.
What are the critical components of a spectrophotometer?
Light source, monochromator, sample holder, and detector.
What is the Beer-Lambert law?
A linear relationship between absorbance and concentration:
𝐴=𝜖⋅𝑐⋅𝑙, where 𝜖 is molar absorptivity, 𝑐 is concentration, and 𝑙 is path length.
What applications use spectrophotometry?
Clinical diagnostics, water quality analysis, and food safety testing.
Explain how wavelength selection is critical in spectrophotometry.
Correct wavelengths ensure specificity for analyte absorption, improving accuracy.
What are the benefits of LED light sources in spectrophotometry?
They offer stable, monochromatic light and lower energy consumption.
Why are optical fibers used in spectrophotometry?
They allow flexible light delivery and collection in compact or inaccessible setups.
How is SpO2 calculated using absorbance?
By comparing the absorbance at red and infrared wavelengths using the ratio of oxy- to deoxyhemoglobin.
Q: How can you calculate the concentration of a target molecule using a standard curve?
A: Use the equation 𝑦 = 𝑚𝑥+𝑏 from the standard curve to solve for 𝑥, where 𝑦 is the measured absorbance.
Q: What physiological parameters can a pulse oximeter measure?
A: Blood oxygen saturation (SpO2) and pulse rate.
Q: How does a miniature spectrophotometer work?
A: It captures the entire light spectrum instantly using a diffraction grating and photodetector array.
Q: How can you calculate the numerical aperture (NA) using θC?
A: NA=sin(𝜃_𝐶)
Q: How can you calculate the critical angle (θC) using refractive indices?
A: 𝜃_𝐶 = arcsin(𝑛_cladding/𝑛_core)
Q: How does a reflection probe deliver excitation light and collect scattered light?
A: The central fiber delivers excitation light, and surrounding fibers collect reflected/scattered light.
Q: How do you calculate absorbance values from voltage outputs?
Use
𝐴=−log_10(Voltage_sample/Voltage_reference)
