How Does A Spectrophotometer Work

How Does A Spectrophotometer Work

In our post spectrometer vs. spectrophotometer, we explained the difference between the two and how they relate to one another. There, we shared that a spectrophotometer can be any number of instruments that measure light. All spectrophotometers use a spectrometer. 

The spectrometer plays a major role in measuring the light, and many other analytical instruments contain spectrometers. The spectrophotometer is the full system that contains a light source along with a means to collect the light for measurement.

What Does a Spectrophotometer Measure?

Spectrophotometers measure the transmission properties of any given material as a function of wavelength. Essentially, spectrophotometers measure light intensity with wavelengths. They also measure electromagnetic intensity at various wavelengths.

Because they measure the frequency emitted by the substance that’s being analyzed, spectrometers do not have their own unit to determine the frequency emitted. Instead, measuring units are based on light absorption – wavelength and light intensity.

The wavelength of light transmittance or absorbance is measured in nanometers. Because of the small size of the length, the human eye cannot accurately detect it, so machinery is required.

The spectrometer is also capable of providing results on light intensity. However, this requires using multiple complex formulas to calculate the object or sample’s transmittance.

Spectrometers measure a wider range than visible light, which represents just a fraction of the wavelengths of light. The full wavelength of light goes from the gamma-ray (10-5 nanometers) to radio waves (1013 nanometers). Radio waves can be thousands of meters long. The gamma-ray is so small that it is not visible to the human eye.

How A Spectrophotometer Works

Let’s first break down all the parts of the instrument, as this makes it easier to understand how everything works together.

  • Light source: This is what provides the wavelengths of light at great intensity. The range spans from near-infrared to inside the ultraviolet range, and it includes the visible light spectrum.
  • Prism: Also known as the diffraction grating, this is what separates the light source into specific parts of the spectrum. When the variable wavelength selector is adjusted, the prism’s position changes so that different wavelengths of light are directed toward the sample compartment that contains the object or sample being analyzed.
  • Variable wavelength selector: This component is on the outside of the instrument and allows the light to be filtered so that it only transmits light at a certain wavelength or range of wavelengths.
  • Sample compartment: Here is where you’ll find the transparent tube, also known as a cuvette, that holds the sample you want to analyze, known as the analyte. The wavelengths you select with the selector pass through the analyte, which are then detected by the photodetector.
  • Photodetector: Light that passes the sample being analyzed hits the photodetector, which is made of semiconducting material. Electrons in the material are excited proportionally to the wavelength that strikes the photodetector.  Increasing the light intensity produces additional electrons, so the signal processor receives a higher current.
  • Display: This component displays the transmittance of the sample. Many models also display the sample’s absorbance, too.

An important part of the entire instrument is the entrance slit because the size of that slit determines the amount of light that can enter and be measured. This affects not only the speed of the spectrometer’s engine but also the optical resolution. The optical resolution is expressed as the full width at half maximum. Smaller slit sizes translate to a better resolution. The slit can be adjusted to allow for more or less light to enter the spectrometer. 

After the light passes through that entry slit, it hits the prism and refracts, then passes through to the sample, which is measured.

Spectrometer vs Spectrophotometer

These terms sound very familiar but have some key differences.  We go into depth on this in our post Spectrometer vs Spectrophotometer, but here is a quick summary explaining the differences.  

A spectrometer is any instrument used to measure the variation of a physical characteristic over a spectrum. These tools are used to collect information about a material based on the amount of infrared, visible, or ultraviolet light it projects.

A spectrophotometer, on the other hand, refers to a number of instruments that measure light. The exact definition varies depending on the area of science or industry. In all situations, the term “photo” is used to indicate that the spectrometer is used to quantitatively measure light intensity with wavelengths. They can also measure the intensity of electromagnetic radiation at numerous wavelengths.

Types of Spectrophotometers

Spectrophotometers are either single-beam or double-beam. Single-beam spectrophotometers measure the light intensity before and after the sample is introduced, while double-beam spectrophotometers compare the intensity of light between the reference light path and the sample that’s being measured. Double-beam models are more accurate because they are not as sensitive to light source fluctuations, but single-beam options have a higher range and are more compact.

Infrared Spectrometer

Infrared spectrometers, sometimes referred to as IR spectrometers, measure vibrations in the interatomic bonds within the sample being tested. When the sample is exposed to infrared wavelengths, the vibrations are measured at different frequencies. This spectrometer can also measure the number of absorbing molecules.

Infrared spectrometers can identify and study chemicals in gas, solid, or liquid form. It is useful for forensic analysis, organic and inorganic chemistry, microelectronics, manufacturing, art history, and various other applications.

Raman Spectrometer

Raman spectrometers are most often used in chemistry to provide the structural fingerprint to identify molecules. This type of spectroscopy relies on inelastic scattering of photons. It uses a source of monochromatic light, typically from a laser. Generally, it’s in the visible light, near-infrared, or near-ultraviolet spectrum, though it’s also possible to use x-rays. The laser interacts with excitations within the sample, which shifts the energy either up or down. That shift provides information about the vibrational modes, similar to the information infrared spectroscopy offers.

UV-Vis-NIR Spectrometer

UV-Visible spectroscopy exposes the sample to ultraviolet light, which excites the electrons upon absorbance of the light energy. The absorbance is measured based on how excited the electrons become. This type of spectroscopy is commonly used to research the chemical bonding of molecules in the sample material.

Near IR spectroscopy is based on the absorption of electromagnetic radiation at wavelengths from 780 to 2,400 nanometers. The light interacts with the sample and then the detector measures the transmittance and absorbance. Near IR spectroscopy has a wide range of applications, including, neonatal research, blood sugar, functional neuroimaging, urology, ergonomics, atmospheric chemistry, and more.

X-Ray Spectrometer

X-Ray spectrometers excite the inner electrons of the sample. When the excited electrons fall into the empty space generated as a result of energy absorption, x-rays are procured.

Regardless of the type, make, or model of spectrophotometer your lab requires, Excedr can help you get what you need.

Leasing vs. Buying A Spectrophotometer

Spectrophotometers are expensive pieces of equipment. No matter how established your lab is, purchasing either new or used can take a big bite out of your budget. Leasing your lab equipment, on the other hand, enables you to acquire equipment you need at a much more affordable price point up front. And because Excedr takes care of all the necessary equipment maintenance and upkeep, you can focus more of your time and effort on the tasks that really matter.