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Abstract
This research review analyzes five articles using the theoretical parameters of UV-vis spectrometry. The technique gains its significance as an instrumental analysis tool that determines the micro and semi-micro amounts of analytes within a specimen while aligning to the wavelength. The derivative spectra of UV-vis spectrometry on these articles are discussed from the first to the fifth article. This includes green tea-samples, wine vinegar, meteorite samples, whisky brands and counterfeit identification, and counterfeit alcohol beverages identification. Thus, offering a vital understanding of the application and limitations of this methodology for chemical analysis. Methods and measurement techniques for acquiring UV-vis spectrometry are described. The magnitude of polynomial fit on UV-vis spectrometry smoothness is discussed. Uses of UV-vis spectrometry in determining the different multi- and single components of these areas analysis are highlighted. Also, UV-vis spectrometry will examine types of species absorbed within the UV-vis zone for the various subjects , where these comprises the organic molecules, species involved with charge transfer complexes and inorganic salts with ions of the transition elements. A summary is made for the research work touching significant details. Lastly, the advantages and disadvantages of the subjects within the five articles work using UV-VIS spectrometry are discussed as well under the critical discussion.
Keywords: UV-vis Spectrophotometer derivative; Measurement techniques; UV-vis spectrometry critical discussion.
UV-VIS spectrometry is among the ancient instrumental methodologies of analysis that covers various ideal techniques to identify the micro and semi-micro samples within an analyte. The technique was first pioneered in the 1950s, however due to the complications involved in generating the derivative spectra through the Uv-visible spectroscopy the technique was used less. The limitation was dominant during the 1970s when computers provided the spectra with more elaborate, rapid, specific and reproducible features. The determinations help in the field of clinical laboratories, research, chemical analysis and industry. Thus, it is vital to study the origin of the UV-VIS spectrum and its features.
UV-VIS spectroscopy
It represents a spectroscopic approach, which draws a variation in uv-visible absoprtion, IR and fluorescence spectrometry. This derivative technique focuses on various objectives within the analytical chemistry area.
-spectral differentiation
-quantitative analysis
-spectral resolution enhancement
Spectral differentiation – is a qualitative technique that differentiates small changes between almost identical spectra.
Quantitative resolution enhancement – it covers the multi-component examination and corrects the unnecessary absorption within a background. Derivative spectroscopy technique creates the start of the distinction or resolution for the overlapping bands, and the important features of derivative procedure are that wide bands undergo suppression with regard to sharp bands relativity.
Spectral resolution enhancement – in this area, the overlapping bands are resolved to enable the simple estimation of the number of wavelengths and bands.
Techniques of measurement for UV-VIS spectrometry
The zero order differentiation spectrum of elements combination demonstrates the manner for any derivative spectrum order. Several methods are used to discriminate a spectrum viz., through numerical or analog method, where the spectral distinction can be intentional either registered within memory of a computer or graphically represented on a paper (5). The spectral value determination is attained by three techniques namely zero crossing , numeric measurement and graphic measurement.
Zero crossing
It measures the derivative spectra at a specific wavelength, at a point which the derivative crosses the zero line. Disruption of one part in the measurement of other element can be removed by this technique.
Numeric measurement
This technique applies groups of points where the spectral values are performed by approximating the derivative value at a particular wavelength. It provides derivatives via spectral variation applying appropriate numerical algorithm.
Graphic measurement
This is a conceptual method that determines the derivative spectra from a paper. This technique is manual thus can offer inaccurate outcomes since the value calculated numerically can be dismissed or abolished beyond the restriction point.
Uv-vis spectrometry
Within the quantitative analysis, derivative spectra extend the distinction between spectra to create overlapping bands. The digital methodology preferably used to obtain the derivative algorithm is called the Savitzky-Golay. It entails plotting the absorbance vs. wavelength rate of change. Also, derivative spectrum of UV-VIS spectrometry can be generated by different experimental methodologies; the differentiation can be performed numerically even when the spectrum has been computerized or noted digitally. If the spectrum undergoes scanning at a continuous rate then a real time recording of the spectra can be realized through the wavelength modulation or time derivative of the spectrum. A wavelength modulation tool will be used to note the derivative spectra, where a radiation beam is different in wavelength at a change of (1-2nm) and the two readings variations is noted, a computerized approach will be applied in obtaining the derivative curves.
For the second and fourth order derivative curves, quantitatively the peak heights are determined from the short-wave and long-wave peak satellite. The magnitude of derivative spectra adjusts with the satellite peaks presence. Contrastingly, the second derivative spectra are displayed by the troughs and two sharp peaks presence. Thirdly, the solvents possess astonishing influence over peaks. From the solvents polarity, troughs and peaks change either to longer or shorter wavelength (fig. 1).
Procedure of acquiring the derivative spectra
UV-VIS spectrometry achieves normal or zero conversion for spectrum orders to the first, next or greater derivative spectrum. It produces considerable alterations within shape of the realized derivative. Essential selection of this derivative order would provide necessary isolation of overlapped signals. Procedure such as width, height and the distance existing between maxima within basic spectrum is attained through control derivative order. Thus, achieving the broad spectrum for the bands, there is needed to apply low orders, while higher orders necessitate narrow spectral bands. Another ideal absorption band that gives clarity in idea for the transformation happening within the derivative spectra is the Gaussian band. Consequently, plotting absorbance versus the wavelength would generate a graph that displays a peak with minima and maxima that should pass via zero upon the ordinate [10](fig 2).
Zero order (UV-VIS spectrum)
This derivative order represents the first criterion of providing further derivatives. For instance, zero th order spectrum can provide the n th order spectral derivative. Zero th order in derivative spectroscopy represents the standard absorption spectrum. This means that the first, second, third and even fourth orders can be directly generated from the order spectrum (zero th). Derivatives order increase enhances the sensitivity of measurement. When a spectrum is highlighted as absorbance (A) being a function of wavelength (λ), the spectra derivative will be expressed as,
A= f(λ)
Derivative spectrum of the first order
This is the spectra generated by derivation of the zero order one in a spectrum. It also represents absorbance plot change with wavelengh against wavelength. It is expressed as.
dA / d λ= f (λ)
This order is more complex than the zero order spectrum even at its derivatized form. The spectrum of the first order goes via the absorbance band’s (λ) max. This absorbance band from the first order will display particular negative and positive bands with minima and maxima. When the spectrum is scanned with constant and minimum difference between two wavelengths, the dual-wavelength spectrophotometer acquires first-derivative spectra.
Derivative spectrum of the second order
This order spectrum is produced from derivatizing of the absorbance order twice. It is a representation of absorption curvature spectrum versus the wavelength (16). The order is given by the equation below.
d2A / d λ2= f ‘(λ)
This order possesses a direct correlation with concentration. This means it is directly proportional, where d2A / d λ2 have to be extensive, the larger the ratio the higher the sensitivity. This technique is essential for acquiring gas and atomic molecular spectra.
Derivative spectrum of the third order
This order unlike the second order demonstrates a disperse function towards an original curve [11].
d3A / d λ3= f ”'(λ)
Fourth spectrum
This represents the inversion of the second order spectrum with a sharper central point than the initial band. Narrow bands are specifically measured by the Uv-high pressure. It is expressed as shown below.
d4A / d λ4= f ”(λ)
Polynomial degree
This feature possesses a higher effect on various polynomial peaks rather than the derivation shape. Polynomial degree scope is low, and applies a half-width differentiation spectra by less degree polynomials, and spectra of small half-width contains higher degree polynomials. As such, the distorted degree spectrum originates from improper polynomial degree. In circumstances of multi-element analysis, the assayed compounds spectral differences and their selective measurement can be improved by using distinct polynomial degrees.
Spectra smoothing
Smoothing operation is undertaken on UV-VIS spectra separately on every data row and its correspondent variables. Derivative spectrum can be changed when a higher degree smoothing is employed, thus the need to take precaution. Smoothing impact relies on tow variables namely; ratio of smoothing; and smoothing frequency. This is the ratio of the smoothed peak compared to the data points number (M).
Tabulated Critical analysis of the papers
Papers
Analysis review
Whisky brands and counterfeit identification
Methodology: Samples were investigated within triplicate on UV–Vis spectrometry. Absorbance data were obtained at 1 nm resolution between 190 and 1100 nm. This was transferred to the spreadsheet via the UV–Visible Chemstation software.
Inference: good outcomes from the authentic samples but two false positive misclassification samples were realized.
The UV–Vis spectroscopy method in collaboration with PLS-DA demonstrated to be an effective technique for the separation of the seven whisky brands. The Spectra were obtained only requiring no sample preparation and a small volume. The recommended discrimination techniques shoed a great promise and a small quantity of outliers within modeled brands/predefined and non-occurrence of over fitting were displayed.
However, there was incorrect results identification PLS-DA in brands counterfeit. This can be corrected by observing all outcomes (the Q residuals, T2 , and estimates limit for the class numbers )not just the minimum value for delimitation.
Meteorite samples
Methodology: 30 meteorite samples of reflectance spectra were measured using UV-vis-NIR spectrometer (laboratory spectrometer).
Inference: The determination resulted in 30 spectra from the samples as highlighted in fig 5-7.
The meteorites have a link to the asteroids. The spectral behavior shown by these samples are identical, but not directly the similar. Differences were recorded in the absolute reflectance level, within the absorption strength features, and upon the spectral slope. These variations are a factor of surface characteristics.
In the future, PCA tool can be used to enhance the performance of Uv-vis spectrometer by revealing the hidden similarities and enable one to link the meteorites counterparts making them more reliable.
Counterfeit alcoholic beverages identification
Methodology: This technique applied PCM spectral data matrix 306 of dimension. This kind of approach helps with outsourcing surges minimizes the data range and classifies data to simplify the spectral study subject.
When the data range is minimized, the approach solves the counterfeit products issue by measuring the similarity degree of the samples by applying the cluster analysis.
It also applies the Gaussian multivariate (for classification purposes) and hierarchical (g legal and counterfeit clustering) analyses.
Inference: this technique is more viable since it can simulate over 153 samples. The application of word method in this approach makes it efficient and reliable to reproduce more than what is offered by other spectral methods.
This technique applying the Uv-vis spectrometer has 100% accuracy in identifying counterfeit products. Thus, applying this methodology of Uv-vis spectrometry can enable implementable objective of minimizing the fatality rates connected to alcohol consumption to be performed.
green tea samples
Methodology: sample measurement under triplicates. It used UV–vis spectroscopy with ultra violet and visible spectroscopy, infrared spectroscopy and Chemometric analysis.
The Uv-spectroscopy data zone were under PCA
.The tested samples were successfully discriminated into clusters using the PC1 and PC2.
PC1 accounted for 73 % while PC2 at 21% of the total variance data at 94%.
Secondly, performing the PLS-DA enables the reinforcement of isolation between the green tea samples acquired from distinct localities.
Inference: UV spectroscopic effectively assisted in the discrimination and authentication of green tea samples sourced from different regions ( East and south Asian areas). This was achieved through data chemo metrics and combination.
This would be vital in promoting quality herbal drug optimization depending on the active predominant constituents.
Wine vinegars discrimination by UV–vis spectroscopy
Methodology: 50 PDO wine vinegars were analyzed using PCA (Principal Component Analysis), HCM (Hierarchical Classification Model ) for two classes classification and PLS-DA spectroscopy (for sample classification) techniques.
Bootstrapping was used to provide consistent and reliable confidence limits for the methodology.
Inference: UV-VIS spectrometer using the above approaches shows that the Uv-spectra region can be delineated using spectra of 300nm, and the visible area between 500 and 600 nm. Thus, explaining the aged vinegars variation. Also, it highlights the intensity and displacement of the Uv regions. Spectral range was also affirmed by UV –VIS absorbance bands in this methodology.
This technique is simple and fast. Although Uv-vis has some unspecific data due to aging compounds of unspecified origin, UV-VIS differentiated the wine vinegar types and origin using this technique.
Therefore, using the Uv-vis spectra together with other systems like SIMCA and PLS-DA can lead to creation of a tool, which offers fast and easy differentiation for the separation of such products in the future.
Summary
From the five articles reviewed under this work, UV-VIS spectrometry is demonstrated as an instrumental method. It offers ideal techniques for the determination of various quantities (micro or semi-micro) within different analytes. This spectrum is triggered by the transitions happening in the electronic energy capacities of the various absorbing specimens. For samples in the liquid state , the spectra takes the shape of a continuous and smooth absorption peak, for gaseous state samples, the spectrum comprises a closely compact lines. These aspects occur because of the interactions and collisions within the solvent molecules and absorbing species. As discussed previously in the introduction, a model UV-VIS spectrum is represented by the wavelength against absorption plot, and characterized by the intensity and absorption band maximum position (λmax).
The five articles covered wine vinegars, meteorites, counterfeit alcoholic beverages, whisky brands and green tea samples discrimination. It is imperative to note that UV-VIS region absorbs categories of species. These include species that are characterized by charge transfer complexes formation, the inorganic salts and the organic molecules. The correlation of the absorption of these elements together with absorbing component path length and concentration relies on Beer-Lambert’s law. This hypothesis is the main fundamental for quantitative measurements undertaken on the analytes highlighted above. Additionally, factors such as electrolytes concentration, association and dissociation possibility, ph and the wavelength can as well result in deviation from this law.
From the review performed on the articles. It is apparent that UV-VIS has five primary components. They include: sample holder, radiation source, detector, and output device and signal processing, and wavelength selector. Uv- region uses a deuterium lamp as its source, while employing the tungsten lamp as the visible range. The selection of the wavelength is attained through filters absorption within low cost parameters for the visible region. However, contemporary instruments use suitable monochromators, where samples are taken using quartz cuvettes. The transmitted radiation is carried out by applying a phototube. Currently, the machines created uses diode arrays to detect samples. These elements are organized into diode array spectrometry tools, single and the double beams.
Advantages
UV-VIS spectrometry has been instrumental in enhancing selectivity and sensitivity. The methodology has multiple merits to its name. First, it helps detect a single or more elements within a solution, and determine these samples concentration. Secondly, it assists with generating structural data about substances; especially the organic compounds, thus helping to highlight a molecule identity. Thirdly, in qualitative analysis, UV-VIS spectrometry help detect unsaturation within a molecule. Fourth, alterations in spectra from the ph changes in a solution can generate essential data on an analyte’s nature. Fifth, the broad absorbance in UV-VIS spectroscopy supports the specific wavelength idea at the maximum spectrum of the technique.
Finally, the quantitative determination of UV-VIS spectrometry is useful in discriminating the biochemical, organic and inorganic species. This has been evident throughput the five articles analyses. These determinations have followed four procedures of identification, wavelength selection, factors influencing the absorbance and Beer and Lambert’s law validation.
Disadvantages of UV-VIS spectrometry
Although it is a sensitive technique, UV-VIS spectrometry is highly affected by various aspects. First, the stray light of this method can lead to faulty design of the equipment. Second, the quality of the detector circuit and electronic circuit design can influence the noise quantity coupled into the signal measurement system, thus resulting in inaccuracy, and sensitivity reduction on the instrument. Third, the similarity in shape for the zero order spectrum and derivative spectra can strongly affect the derivative spectrum. Fourth, UV-VIS spectrometry is confined to a specific system, thus causing limited usages because it is not easy to reproduce. Lastly, the inability to reproduce can change outcomes, leading to various spectrophotometers being applied for zero order spectra; thus offering same outcomes but different derivatization display.
Reference
De Caro, Cosimo & Haller, Claudia. (2015). UV/VIS Spectrophotometry – Fundamentals and Applications.
Hussain, Alaa. (2019). UV-VISIBLE SPECTROMETRY.
https://scielo.conicyt.cl/scielo.php?script=sci_arttext&pid=S0717-97072018000304126 (sample format)