Selection of Cuvette in UV-Vis Spectroscopy

Absorbance studies have become a base for the examination of numerous particles and solutions. For this to happen, light needs to be passed in a defined format. The format is provided by cuvette. The selection of the cuvette is a significant element in the measurement of absorbance in the UV-Vis spectrophotometer as solutions are placed inside the cuvette for analysis. Also, the selection is a delicate task as there are varieties of cuvettes available. This blog gives an introduction to the cuvette. Then, types of cuvette are provided. It also speaks about the way of the usage of the cuvette. Subsequently, it discusses how different factors affect the selection of cuvette in the UV-Vis spectroscopy system. 

What is Cuvette?

• The cuvette is rectangular test tubes. They are used to hold aqueous solutions like normal test tubes. Normal test tubes are useful in chemical reactions. In contrast, cuvettes are used in UV-Vis spectrophotometer or fluorometer for the measurement of transmittance or absorbance of radiation at a particular wavelength. 
• The solution needs to be placed into the light path of a photometer. Cuvette serves the purpose. Mostly, 80% volume of cuvette is filled. But the filling volume varies for different cuvettes. The sample is directly injected into the cuvette using a micropipette and the cuvette is ready for use. 

Other Names for Cuvette

Cuvettes are called by various different names. They are as follows:

• Cells
• Cuvette cell
• Sample cuvette
• Capillaries
• Quartz cell
• Spectrophotometer cuvette
• Absorption cell

Types of Cuvette and Compatible Wavelengths 

. The popular cuvette materials are as follows:

• Pyrex glass or optical glass
• IR quartz
• UV quartz
• Sapphire

• Glass cuvettes are made from polystyrene, PMMA, and glass. These materials are good for use in the visible radiation range, having a modest transmission range from 340 nm - 2500 nm. In addition, most applications come under this range only. If low money is allocated for buying a cuvette, then it is better to go for the purchase of an optical glass cuvette as it is a cheaper one. 
• UV quartz is a notch above the optical glass and is a special kind of plastic. It is costlier than optical glass, but it is equipped with an extended transmission range from 190 nm - 2500 nm, which is in the UV spectroscopy range. Thus, for measurement of UV spectra, it is compulsorily needed.
• IR quartz is one of the better choices for UV-Vis experiments. It gives a transmission range of 220 nm - 3500 nm. In addition to the IR region, some of the UV regions can also be covered.
• Sapphire cuvette is super hard, making it hard to scratch. In fact, it is resistant to damage caused by electricity, heat, and chemical corrosion. It has one of the widest transmission ranges of 250 nm - 5000 nm. Since the optical range is broader and the quality of the material is higher, it is one of the most expensive materials.

Selection of Cuvette in Ultraviolet Visible Spectroscopy

When looking for a measurement of absorbance in a UV-Vis spectrophotometer, selecting a proper cuvette is an essential thing as each cuvette material has its own pros and cons. Depending on the application of UV spectroscopy, the material is chosen.

• Each cuvette has a uv-vis absorbance cut-off. Below the cut-off value, the solvent itself absorbs the light.
• For UV-based absorbance, it is expected to use only quartz cuvette. However, for measurement of absorbance in the visible region, glass, plastic, and quartz cuvettes are acceptable.
• For experiments requiring high purity, disposable plastic cuvettes are preferable as they minimize the risk of contamination. For an aqueous solution, any cuvette can be used. But for using organic solvents, glass cuvettes are selected as they display higher resistance to solvents when compared to plastic ones.
• Plastic cuvettes can be reused only a few times, but it is cost-effective. Since glass and quartz are reusable materials, they can be washed and maintained well, so that they can be reused for a longer time. Quartz has the capability of more light transmission and is more transparent. So, it is used for sensitive experiments. Besides, quartz is more temperature resistant. At the same time, glass and plastic cuvettes are not ideal for concentration and purity measurements.  
• Cuvettes are polished on two or four sides. The polished side allows maximum transmittance of light. Since light passes through the sample in a linear fashion, polishing two sides is enough. For fluorescence and scattering experiments, the signal is received at 90° to the light, hence all four sides require to be polished.

Dimensions, Usage & Maintenance 

• The dimension of a commonly used cuvette is 12.5 mm*12.5 mm. Sample volumes can be in the range of microliters to milliliters. Generally, the heights of light paths are 8.5 mm and 15 mm. The path length is 10 mm.
• The contact area present between the wall of the cuvette and the shaft holding the cuvette needs to be as large as possible, to facilitate efficient temperature and heating control over the sample.
• The Cuvette is closed at one end. It should be clear without impurities as impurities can affect the reading of a spectrophotometer. It is open to the atmosphere at the top and sealed at the bottom. The chosen cuvette should be transparent. Cuvettes should be placed in a dark area chamber for measurement. The chamber can have rotating carousels, which are able to hold many cuvettes.
• Cuvettes are fragile and expensive. Hence, they should be properly used. Cuvettes should not be scratched. They should not be stored in wire racks. In addition, they should not be cleaned with abrasives or brushes. Samples should not be present in the cuvette for a longer period of time. They should be washed immediately after usage. Disposable plastic cuvettes are cheaper ones. 

Things to Remember

There are some precautions that need to be taken while using cuvette for measurement of absorbance.

• Initially calibration of UV spectrophotometer should be done with blank measurement using cuvette.
• If higher dilution in a cuvette compartment is done, there will be a decrease in signal to noise ratio.
• Measurement below 190 nm should be done with oxygen free nitrogen purging in the sample compartment and if below 185 nm is needed, vacuum should be maintained in the sample chamber. 

Conclusion

Cuvette is able to influence the result of absorbance directly or indirectly. Choosing an appropriate cuvette for the measurement of absorbance and transmittance in UV-Vis spectroscopy requires utmost care. The quality of the cuvette should not be compromised, otherwise experimental results will be impacted. Additionally, care should be taken of obtaining good cuvettes within a stipulated budget.