This presentation gives a overview about the microvolume UV/VIS spectroscopy. Instrumentation , working ,merits, demerits, cleaning of the sample platform. mainly explains about the measurement of sample using nano or micro volume samples.

1. MICROVOLUME UV/VIS SPECTROSCOPY
PRESENTED BY
SURIYAPRIYA.K
1st yr. M.PHARM
DEPARTMENT OF PHARMACEUTICAL ANALYSIS
KMCH COLLEGE OF PHARMACY
COIMBATORE

2. UV-VIS SPECTROSCOPY (OR SPECTROPHOTOMETRY)
It is a quantitative technique used to measure how much a chemical substance absorbs light.
This is done by measuring the intensity of light that passes through a sample with respect to the
intensity of light through a reference sample or blank.
MICROVOLUME UV-VIS SPECTROSCOPY
UV/VIS micro-volume spectrophotometry is a technique used to measure intensity of light with
only one drop of sample. So it is referred as to as low volume ,small volume ,nano
spectrophotometer . A wide range of concentrations is covered by automatically measured .

3. EVOLUTION OF UV /VISIBLE SPECTROPHOTOMETER :
 In 1930s, American government’s interest in measuring vitamin contents using UV/Vis light.
 In 1940 commercial launched UV-Vis spectrophotometer.
 In 1941, Beckman introduces the DU UV-Vis spectrophotometer.
 In 1946, Howard Cary is founded spectrophotometer named as Cary Instrument.
 In 1950, National Technologies Laboratories changes its name to Beckman Instruments.
 In 1953, Bausch & Lomb introduces the SPECTRONIC 20 UV-Vis spectrophotometer.
 In 1954, Applied Physics Corporation launches the Cary 14 spectrophotometer.
 In 1963, JASCO introduces the ORDUV-5 with double-beam UV-Vis capabilities.
 In 1969, Cecil Instruments introduces the CE 21.
 In 1979, Hewlett-Packard launches the first commercially available diode-array spectrophotometer.
 In 1980, Bausch & Lomb introduces the Spectronic 2000 UV-Vis spectrophotometer.
 In 1987, Pye Unicam Corporation. introduces the PU-8700 UV-Vis spectrophotometer.

4.  In 1995, Hewlett-Packard launches the 8453A, diode-array spectrophotometer.
 In 2000, Thermo Scientific introduces the GENESYS 10 instruments .
 In 2002, Varian Inc. releases the 6000i UV-Vis-NIR spectrophotometer.
 In 2003, Thermo Scientific introduces the Evolution 300 spectrophotometer, the first double-
beam xenon lamp-based spectrophotometer.
 In 2004, Shimadzu introduces the SolidSpec-3700/3700DUV series of UV-Vis-NIR
spectrophotometers.
 In 2005, the Nanodrop ND-1000 UV-Vis spectrophotometer .
 In 2008, Shimadzu launches the UV- 1800 compact UV-Vis spectrophotometer.
 In 2010, Thermo Scientific introduces the Evolution 200 Series spectrophotometer.
 In2010, JASCO offers the SAH-769 One Drop.
 In2011, Agilent Technologies releases the Cary 60 UV-Vis spectrophotometer.

5. MICRO-VOLUME UV/VIS SPECTROPHOTOMETER
INTRODUCTION :
 Spectrophotometers are instruments used for measuring radiant or light energy, traditionally
with a quartz cuvette.
 Improvements in technology now permit measuring in capillary-type or sample plate
containers.
 One of such type of instrument is micro volume UV /visible spectrophotometer .
 Microvolume spectrophotometers, also referred to as low volume, small volume or nano-
spectrophotometers.

6. INSTRUMENTATION
LIGHT SOURCE : pulsed xenon flash lamp ,Dedicated xenon flash lamp
SAMPLE CELL : Sample platform ,cuvette
DETECTOR : CCD( charged coupled device), photodiode
READER : Lab X pc software

7. MICRO-VOLUME UV/VIS SPECTROPHOTOMETER
WORKING:
 Mettler-Toledo provides a spectrophotometer capable of performing micro-volume UV/VIS
measurement.
 This instrument is capable of measuring very small volumes and highly concentrated
samples.
 The method is fairly straightforward.
 The sample is pipetted directly onto the measuring platform, without further dilution.
 Therefore, manipulation errors are avoided.
 Moreover, the selection of a specific path length allows for the measurement over a large
concentration range with as little as 1 µL of sample.

8.  The measurements are performed at the micro-volume platform covered by a movable
arm mounted on the top of the instrument.
 Spectrophotometer has both a micro-volume platform as well as a cuvette holder.
 Depending on the selected application, the light can be directed either onto the micro-
volume platform or the 1 cm cuvette.
 The transmitted light is then focused on the grating where diffraction occurs.
 The diffracted light beams of different wavelengths are then directed onto the detector.

9.  When the arm is in open position, the micro-volume platform can be easily accessed with
a pipette from either the left or the right side.
 The curved lid on top of the instrument allows convenient positioning of the operator's
hand to securely guide the pipette tip.
 During measurement, the arm is securely locked to a precisely defined path length and
cannot be opened until the measurement is completed.
 The transmitted light is acquired by a detector charged coupled device .

10. INSTRUMENTATION OF MICROVOLUME UV/VIS SPECTROSCOPY

12. CLEANING OF PLATFORM
 On a micro-volume platform, samples with very small volumes, such as
microliters, can be measured
 An initial cleaning of both measurement surfaces with dH2O is recommended.
 dH2o is recommended for blank measurement in order to avoid interferences
with the measurements
 Do NOT use a spray bottle to apply water or any other liquid to the surface of the
instrument.

13. Additional cleaning recommendations are as
follows:
 Between measurements it is recommended to wipe the sample from both the upper and
lower platforms with a clean, dry, lint-free lab wipe.
 Depending on the sample, a 60% isopropanol solution, ethanol or ultrapure water can be
used to clean the micro-volume platform.
 If necessary, the solvent used to dissolve the sample can be applied.
 When samples dried on the platform, additional cleaning can be performed using 3 µL of
0.5M HCl.
 Finally , If detergents or 100% isopropyl alcohol are used, follow with 3 µL of dH2O as final
cleaning step.

14.  The polished surface allows for droplet application of aqueous solutions with high contact
angle.
 If the droplet flattens out, it is possible that after closing the micro volume arm the
measurement is performed through air, which will lead to erroneous measurement results.
 This is visualized in the following images
 For decontamination, a solution such as 0.5% sodium hypochlorite (commercial bleach
solution 1:10 diluted) can be applied to the micro-volume platform.
 An additional cleansing with deionized water should follow.

15. SPECIFICATIONS OF UV5NANO
SPECTROPHOTOMETER
U

17. THE UV5NANO SPECTROPHOTOMETER

18. TRICKS AND TIPS FOR ANALYSIS:
UV5NANO MICRO-VOLUME
SPECTROPHOTOMETER
 Effectively clean the sample area
 Properly set the pipette
 Improve repeatability
 Pipette smoothly on the platform
 Determine rhythm and speed

19. MERITS
 Accurate Microvolume Measurements
Only 1μL of sample is required for reliable measurements.
 Measure Wide Concentration Ranges Fast
 Double Application Power
The UV5Nano is two instruments in one: you can perform micro-volume or cuvette
measurements.
 Convenient and secure
 Robust and easy to maintain
With no moving parts in the optical set-up, the instrument is robust and easy to maintain.
The flat surface of the micro-volume platform makes cleaning fast and easy.
 Ready to use
The Xenon flash lamp has no warm-up time. Simply power it up and start measuring.

20. DEMERITS
 Light source from xenon flash lamp is expensive.
 Skilled operator is required for handling sample pipette.

21. APPLICATION:
 The capacity to measure volumes in the range of 0.5 to 5 ul,
 Some models can measure even lower.
 These instruments allow for quick and accurate quantification .
 Gives accurate quantification of nucleic acids and protein samples , etc
 Ideal for life science applications such as concentration and purity determinations of DNA, RNA
and protein samples

22. FUTURE OF UV-VIS SPECTROPHOTOMETERS
 Future improvements in UV-Vis spectrophotometers will focus on ease-of-use,
portability, and application-specific instruments.
 UV-Vis analysis of solid samples and materials continues to grow in areas
such as solar cell research, semiconductor products, and coating materials.
 Advances in light sources will provide new developments in conventional
spectrophotometers and handheld UV-Vis instruments.
 Further development in remote sensors will enable more types of samples to
be measured outside the laboratory.

23. REFERENCE
Journal UV/VIS spectroscopy - fundamentals and applications
https://www.labmanager.com/laboratory-technology/evolution-of-uv-vis-spectrophotometers-18481
https://www.mt.com/in/en/home/products/Laboratory_Analytics_Browse/uv-vis-spectrometers/micro-uv-vis-
spectrometers.html
https://youtu.be/8OW6CLknA6Q