The fluorescence spectra of various brands of mango juices that were bought from the local market are displayed in Figure. 1, along with mango pulp from two distinct types (Anwar Ratol and white Chaunsa). Significant emission bands of fluorescence were detected at 445, 469, 522, 545, 580, 634, 674 and 772 nm. These bands were seen in mango juices of different brands as well as in pure mango pulp. Water content is linked to bands at 445 and 469 nm [14]. Beta carotene is identified by the fluorescence emission bands at 525–545 nm [15]. Figure 3 depicted 580 nm band evolution that indicates the addition of tartrazine food color to diluted mango pulp[16]. According to [17], the fluorescence bands at 634, 674 and 772 nm in mango juices indicate the amount of chlorophyll. The most important pigments in mango fruit are beta carotene and chlorophyll, as seen in Fig. 1 which also shows that these bands have a higher relative intensity. All of the mango juices can be compared by considering the relative intensity-based concentration of chlorophyll and carotenoids.
According to [18] when fruit ripens the amount of beta carotene in the peel increases while the amount of chlorophyll drops. Mangoes undergo color changes during ripening because of the breakdown of chlorophyll and the synthesis of pigments called carotenoids. Because mango fruit contains chlorophyll, a powerful antioxidant, it is initially green in color. Depending on the cultivar, the rise in carotenoids causes the color of mango to shift from green to yellow, orange or red. Mangoes undergo changes in texture and flavor as they ripen due to the synthesis and accumulation of sugars, acids and aromatic compounds as the fruit softens[7].
Because of this, the fluorescence emission spectra of ripened mangoes and mango juices exhibit a broad band of beta carotene at 525–545 nm, with varying emission intensities. Figure 1 makes it clear that the majority of mango juices purchased from the market have fluorescence spectra that display a chlorophyll band at 675 nm, indicating that unripened mango pulp is used in the production of commercial juices. Significant variations in the relative intensities of the chlorophyll bands are revealed by their investigation. The usage of unripened pure mango pulp ingredients in Nestle Fruita Vital, Nestle Nesfruta and Shezan mango juices is linked to a significantly greater relative intensity for the chlorophyll band. Conversely, brands such as Fruitien, Fruit Nation, Lush and Quice perfect mango juices exhibit the presence of chlorophyll band, although to a somewhat lesser degree than the aforementioned brands. A lower concentration of unripened mango pulp is suggested by the much lesser relative strength of the chlorophyll band observed in all other mango juices. This could indicate a higher proportion of water content and adulteration of color pigments in their compositions. Mango juices from Fruiten, Fruit Nation and Nestle Fruita Vital exhibit a greater relative intensity band of beta carotene at 545 nm. In comparison to the above-mentioned brands Shezan Fruiticana, Maza, Nestle Nesfruta, Tops Aaam, Quice Perfect, Lush, Real Popular, Tropico and Tops Tangy mango Juices displayed a lower intensity band of beta carotene. The decreased levels of beta carotene in all other mango juices can be the cause of the lowered fluorescence intensity. This indicates that there is less mango pulp and more water in these juices, showing low intensity band of beta carotene.
To examine the impact of water content, various dilutions of mango pulp were made, these dilutions ranged in concentration from 2, 8, 10, 12, 15, 30 and 50%, and the fluorescence emission spectra of these dilutions were recorded (Fig. 2). The main justification for diluting these particular concentrations is that different mango juice brands have varying concentrations listed on their labels. A link between concentrations and fluorescence intensity can be established by comparing the fluorescence emission spectra of different dilutions. According to earlier research, fluorescence intensity and fluorophore’s concentration are directly correlated [19]. There is only one broad band visible in the emission spectrum of the original mango pulp from Anwar Ratol and White Chaunsa at an excitation wavelength of 405 nm.
The spectral signatures of the original mango pulp were significantly different from the samples that were made by diluting pure mango pulp with water. The carotenoid band in these spectra displayed a blue shift, moving from 550 to 525 nm. This shift indicates that the addition of water has altered the qualities or composition of the mango pulp. When comparing the diluted samples to the pure mango pulp spectra, the fluorescence intensity of carotenoid region was higher. On the other hand, the fluorescence emission spectra changed dramatically, displaying extra emissions peaks when the mango pulp was diluted with water. In this diluted situation, the fluorescence intensity starts to increase at the slightly shorter wavelength of 480 nm. Due to the addition of water molecules, there is a shift in the fluorophore environment which in turn causes intermolecular interactions and a shift in the beginning point of fluorescence emission intensity. Due to the presence of water in the samples, additional emission bands at wavelengths of 444 and 467 nm emerged in these diluted samples. This implies that the altered emission bands depend on the water concentration.
According to [20], beta-carotene produces fluorescence maxima about 520–560 nm. The orange hue of mangoes is attributed to beta carotenoids, which are responsible for the 525 and 550 nm peak. Another band that can be seen at wavelengths of 665 and 772 nm is caused by the green pigment chlorophyll, which is frequently found in plants. Figure 2 showed that adding water altered the characteristics of mango pulp and decreased the solute concentration in the diluted samples of mango pulp. This comprises liquid-soluble carbohydrates such as fructose and glucose[21] and flavoring agents. The molecular concentration of these components in mango pulp decreases with increasing dilution of water. The concentration of each ingredient in the solution, including the fluorophores is immediately impacted by dilution. Diluting the solution causes the fluorophores to become less concentrated which may cause the fluorescence intensity to decrease. As a result, the spectra of these diluted samples differ from the original mango pulp. This is frequently explained by the Beer-Lambert rule, which stipulates that absorbance and consequently, fluorescence intensity is exactly proportionate to the concentration[22].
Mango pulp diluted by 50% revealed that the sugar content decreased while the presence of carotenoids increased, leading to high intensity fluorescence spectra with a shift at 525 nm. A declining trend in the relative intensity of emission spectra was seen in the emission spectra of the mango pulp diluted at 30, 15, 12, 10 and 2 percent. This pattern offers a useful instrument for assessing and measuring the amount of water included in mango juices sold in the marketplace. Water content can be assessed by comparing the emission spectra of various commercial mango juices that are relevant to this trend. One important finding from Fig. 2 is that the trend of emission spectra begins to marginally overlap with the spectra of pure mango pulp when the two quantities are identical. Mango pulp spectra that were 50% comparable to the original ones were found to have followed the emission in the 500–750 nm region. Despite having a close resemblance to the original mango pulp, the resultant spectrum displayed a comparatively low intensity and blue shift. Fruits contain a large amount of water when they are fresh, making up a large portion of their weight. According to [23], fresh fruit usually contains 75–95 percent water. Similarly, mangoes themselves contain a significant quantity of water, roughly 75–85 percent of the pulp is water [24]. The molecular interactions between sugars, acid and other substances like water, pectin and cell wall material may be the cause of the physico-chemical property changes[25]. The spectral bands at 444 and 467 nm are directly related to the existence of water content, according to our analysis. Figure 1 made it clear that Premjer young and Popular junior mango juices have quite distinct spectra from other brands. The spectra of both juices show a broad emission band, suggesting that there is a considerable amount of water adulteration in the samples.
Another experiment was created to look into the adulteration of mango juices with artificial or synthetic food coloring using fluorescence spectroscopy. A sample was prepared by mixing artificial color with water at a ratio of 0.5 grams of color to 100 mL of water. In order to create samples with varying levels of color adulteration 84%, 88% and 90% in pure mango pulp that matched with pure Anwar Ratol mango pulp. The fluorescence emission spectra of Anwar Ratol, white Chaunsa mango pulp and 8% diluted mango pulp at an excitation wavelength of 405 nm are displayed in Fig. 3.
The spectra of color-adulterated materials reveal a broad band spectrum with a peak maximum at 545 nm, obtained in the wavelength range of 480 nm to 750 nm. The pattern indicates that the pulp’s fluorescence intensity decreases when the color solution is concentrated. Figure 3 made it clear that the band connected to color addition had a narrower profile, whereas the pure mango pulp band at 545 nm had a wider shape. Mango pulp with color additives showed a spectrum that was almost identical to that of pure mango pulp, with the exception of a little hump that was seen at 580 nm. The original mango pulp does not show a band at 580 nm, as shown in Fig. 3. This band can be used as a biomarker to determine whether commercial mango drinks contain artificial coloring. The eye-catching and vivid hues present in different mango juice brands are produced using approved food coloring chemicals. Every brand makes it quite evident on its labels what kind of food coloring is added, they are usually indicated by codes like E102, E110, or E160a. These codes guarantee precision and conformity to legal requirements, empowering customers to make knowledgeable decisions. It is clear from Fig. 1 that pure mango pulp did not exhibit a peak at 580 nm whereas mango drinks sold in stores, however, have a hump at 580 nm. Among the mango juice options on the market, Fruit Nation and Nestle Fruita Vital have a slight and nearly negligible peak. The relative fluorescence intensity of all other mango juices on the market is far lower, suggesting that they may have been tampered with by adding artificial food coloring and water. Figures 2 and 3 demonstrate how the relative fluorescence intensity is reduced when water and fake food coloring are added.
Heat treatment (HT) is the most widely used technique for increasing the shelf life of juice because it effectively gets rid of dangerous germs and reduces microbiological deterioration [26]. Juice can lose some of its nutritional components when heated because of a number of chemical and physical processes. The impact of varying temperatures on diluted mango pulp is illustrated in Fig. 4. Water, sugar and color additives are added to pure mango pulp before it is heated in the production of commercial mango juices. 8% concentration of pure mango pulp was heated at four different temperatures starting at 60oC. When compared to pure mango pulp, diluted mango pulp (8%) displayed a distinct spectral behavior, with a few additional bands appearing around 445, 467, 525, 550, 665, 743 and 772 nm. This indicates that the molecular makeup of mango juices can be significantly impacted by the amount of water present in commercial juices. The high concentration of conjugated carbon-carbon double bonds present in carotenoids is what gives numerous fruits, vegetables and other natural sources their rich yellow, orange and red colors [8]. Heating diluted mango juice causes an increase in relative fluorescence emission intensity[27]. Thermal treatment can cause oxidation, which can alter the beta-carotene molecule. Oxidation is one of the main reasons food deteriorates and has a reduced shelf life. Furthermore, oxidation results in reactive and hazardous substances that put consumers in danger in addition to reducing the nutritional value (loss of vitamins and essential fatty acids) and flavor of food [28]. These structural differences can affect the electronic transitions and energy levels that result in fluorescence. If conjugated double bonds or other chromophoric groups are altered by structural alterations, the fluorescence properties could be impacted[29].
Some new fluorescent byproducts may arise during the oxidation of beta-carotene in mango juices. These byproducts may have unique fluorescence qualities that differ from beta-carotene’s. Therefore, if these byproducts are more prevalent generally, they may help to raise the fluorescence intensity overall. The evolution of oxidative products, which are visible from the rise in the relative emission intensity of the heated samples, may be the cause. In diluted samples, the blue shifting of the band from 550 to 525 nm indicates a change in the molecular composition of carotenoids following the addition of water. Figure 2 further shows that the carotenoids in these diluted samples develop at 525 nm because there is only 8% mango pulp and 92% water content. Additionally, heating samples of diluted mango pulp causes noticeable variations in the relative strength of carotenoid band at 525. Furthermore, there are noticeable changes in the relative strength of the carotenoid band at 525 nm with a rising trend from 60 to 90oC when diluted mango pulp samples are heated. As temperature rose, beta carotene’s fluorescence emission intensity increased (Fig. 4). The research reported that all-trans-carotene concurrently diminishes and cis isomers rise with thermal processing of mangoes[30]. When carotenoid molecules are exposed to light, oxygen and high temperatures, the existence of cis-isomers becomes more evident[29]. Mango pulp contains a variety of chemicals, including flavonoids and carotenoids which in some situations can exhibit fluorescence qualities. These emission spectra of these components can change depending on pH, temperature and the presence of other molecules. The majority of commercial samples include different concentrations of mango pulp added. Adding mango juice up to 30% showed an increase in fluorescence intensity but adding 50% of mango juice showed a possible decrease that could be caused by fluorescent molecule concentration quenching. The emission spectra of some substances may change as a result of reactions that modify their fluorescence characteristics or break down the luminous molecules. Oxidation which happens when oxygen is introduced to unsaturated fatty acid double bonds is accelerated by heating. According to [31], hydro-peroxides are produced and then decompose into aldehydes and ketones. Processed mango products may also lose part of their aesthetic appeal.
PCA is a technique that is used to categorize complicated spectral characteristics that are difficult to discern visually. By using an orthogonal transformation, it converts correlated variables into uncorrelated principal components [32]. When presented on scatter plots, these components typically two or three capture the majority of the volatility in the data and aid in distinguishing between various datasets. PCA is a data-driven method that helps with complex spectral data analysis and classification by revealing hidden patterns. A scatter plot depicting the reduced-dimensional space produced by PCA is shown in Fig. 5. Every data point on the plot represents a fluorescent chemical and its placements are based on its coordinates on the first two main components (PC1 and PC2), which show different clusters of data points. Compounds that show comparable fluorescence patterns are grouped together and called clusters.
Figure 5 shows how Principal Component Analysis (PCA) was used to classify different mango juice brands into two distinct clusters and some liquids scattered. The juice brands Alita, Funtime, Rock roll, Cooler, Premier youth, Fruitaa, Benz, Golden day, Slice, Popular junior, Tops tangy, Real popular, Quice perfect, White Chaunsa, Anwar Ratol, Tropico, Refresh and 8% mango pulp are seen to be grouped together on the negative side of the PCA plot. This primary set of mango juices can be further divided into two groups: Alita, Funtime, Rock roll, Cooler, Premier youth, Fruitaa, Benz, Golden day, Slice and Popular junior made up group one and second group includes Tops tangy, Real popular, Quice perfect, Tropico, Refresh, white Chaunsa, Anwar Ratol and 8 percent mango pulp. On the other hand Lush, Fruiticana, Maza and Tops Aaam mango juice brands are grouped together on the positive side of the PCA plot. On the positive side of the scatter plot, it is evident that the mango juices Shezan, Nestle nesfruta, Fruitien, Fruit nation and Nestle fruita vital cluster themselves far apart from the other juices, indicating a significantly distinct chemical composition. Group 1 mango juices such as Alita, Funtime, Rock roll, Cooler, Premier youth, Fruitaa, Benz, Golden day, Slice and Popular junior, had a high-water content and artificial coloring added in addition to having a low mango pulp level. It indicates the reason that these juices stand out from other juices is that their quality is not very good. 8% percent of the original mango pulp, white Chaunsa and Anwar Ratol as well as Tops Tangy, Real Popular, Perfect Quice, Tropico and Refresh were grouped together in the second group. Shezan, Nestle Nesfruta, Fruitien, Fruit Nation and Nestle Fruita Vital mango juices are located on the positive side of the scattered plot, somewhat apart from each other and the other juices. Because these mango juices include 30, 08, 30, 32 and 50% of the original mango pulp as stated on their labels, they are dispersed apart from one another and exhibit this scattering pattern. The dispersion pattern of white Chaunsa and Anwar Ratol, the original mango pulp juices, further confirms the existence of mango pulp. It is evident that the juices with higher water contents dispersed themselves starting on the plot’s negative side and moving toward its positive side. While juices with very little water content like Shezan, Nestle Nesfruta, Fruitien, Fruit Nation and Nestle fruita vital lie on the very positive side of the scatter plot, juices with more water content like Alita, Funtime, Rock roll, Cooler, Premier youth, Fruitaa, Benz, Golden day, Slice and Popular junior mango juices lie very close to the negative side.
The original mango pulps were located somewhere in the middle of these two varieties of mango juices with the second group. The commercial juices in this group have a different water content than the pure pulp of white Chaunsa and Anwar Ratol as seen by their labels and outward appearance. Figure 1 further illustrates how similar the emission spectra of these juices are to those of pure mango pulp with both having the same band location at 525 nm. Carotenoids, which show a constant band location throughout all the mango juices in group two are linked to the substantial broad band seen at 525 nm in the spectra of pure mango pulp from white Chaunsa and Anwar Ratol. B-carotene, usually referred to by its European designation E 160 is being utilized as a food pigment. The food industry’s propensity to employ natural coloring chemicals rather than synthetic ones is driving up its use[33]. The presence of water contents in these mango juices may be the reason why pure mango pulp is grouped with these groups. Figure 5 makes it evident that loading vectors, which are based on two primary fluorescence emission bands, explain how these various clusters are classified. One is attributed to carotenoids at 545 nm and the other is a chlorophyll band present at 675 nm. This indicates that PCA used the fundamentals of carotenoids and chlorophyll to describe various brands of mango juice.