Near-Infrared Diffuse Reflection Analysis of Fruit Using the Vivo Direct Illuminated Reflection Stage and NIRQuest256-2.5 Extended Range NIR Spectrometer
The quality of fruit and produce cannot be judged by appearance alone. A colorful mango or vibrant avocado must do more than add a splash of color to the plate. Taste and freshness are critical. Since it is not possible to determine fruit quality by tasting each individual piece, an objective, nondestructive measurement is needed to determine the quality of the fruit hidden beneath the peel. In this application note, near-infrared (NIR) diffuse reflection spectra are measured for avocados and mangoes to demonstrate the power of NIR measurements for the noninvasive assessment of fruit quality.
Consumers use many techniques to assess fruit quality including smell, firmness, sound, appearance and even intuition. Everyone has their own special approach, with mixed results. While these qualitative techniques are sufficient for consumers, commercial fruit growers and packers require a quantitative approach to determining fruit quality to ensure customer satisfaction and retain or expand market share (consider that annual worldwide production of mangoes alone exceeds 62 million metric tons). Determination of critical quality parameters such as sugar, starch and moisture content requires a rapid, noninvasive, online measurement to test fruit prior to picking or packaging. NIR spectroscopy meets these requirements.
NIR spectroscopy has been used since the 1970s for the analysis of agricultural products. Spectral data for NIR light reflected from an agricultural sample like grain or produce is acquired and compared with a calibration model generated from spectral data acquired for samples with known levels of the constituents of interest. For fruit or produce covered by a peel, the longer wavelengths used for NIR analysis are weakly absorbed so they pass through the peel, enabling sampling of the fruit pulp beneath. The measurement of NIR reflection is rapid (no sample preparation is required), quantitative (with the assistance of carefully constructed calibration models) and nondestructive. Since its initial use as an industrial tool in agriculture, NIR spectroscopy has grown significantly in areas ranging from materials analysis to pharmaceutical monitoring and medical diagnostics.
The NIR region extends from 780-2500 nm. Absorption of light in this region causes molecules to vibrate. These molecular vibrations result in spectral data with features dependent on the chemical composition of the sample. In the case of agricultural samples, the NIR spectra are typically composed of broad peaks due to overlapping absorptions caused by overtones and combinations of vibrational modes of organic functional groups like C-H, O-H and N-H chemical bonds. The NIR spectrum provides a snapshot of the sample with information for multiple components available in a single NIR spectrum. These characteristics and others make modern NIR spectroscopy instrumentation ideal for online monitoring and process control.
Starch and sugar (primarily fructose, glucose and sucrose) are commonly measured to determine fruit maturity and quality. While the peaks for these constituents are located near one another, starch has some specific wavelengths that enable construction of a multi-parametric model for determination of fruit quality based on starch components. An extended range NIR spectrometer like the NIRQuest256-2.5 is a great option for these measurements because it can detect critical starch peaks near 1722 nm, 2100 nm and 2139 nm, as well as sugar peaks that occur primarily between 900-1200 nm (some peaks also occur >2100 nm). The NIRQuest256-2.5 enables detection of all these wavelengths in a single spectrum.
In addition to the spectrometer, a bright light source like our Vivo Light Source is necessary. Vivo has four powerful tungsten halogen bulbs and fibers that transmit light efficiently for effective NIR measurements of fruit. Since much of the light will scatter off the surface of the fruit, a large core diameter fiber (600 microns) is recommended for these measurements to increase throughput and improve sensitivity.
Sampling configuration is critical for these measurements. In addition to the light lost by scattering off the surface of the fruit, water in the fruit will absorb NIR wavelengths. In addition, the constituents of fruit (or any natural or agricultural products) are not uniformly distributed within the sample. Sampling over a large surface area of the fruit is recommended to provide an average value for the constituents in the fruit. The Vivo light source with its large illumination area is a great option for sample illumination when testing fruit and produce.
While the results reported here are qualitative, a carefully constructed chemometrics model is required for a multi-parameter, quantitative assessment of fruit quality. With a good set of reference spectra and PLS (partial least squares) modeling, it is possible to develop a calibration model that can be used to measure multiple fruit parameters (sugar, starch and other fruit constituents) for the prediction of fruit quality. The ability to quantitatively measure multiple parameters simultaneously makes NIR spectroscopy a powerful tool for the agricultural industry.
NIR spectra were measured for avocados and mangoes using the NIRQuest256-2.5 extended range NIR spectrometer (900-2500 nm) and Vivo direct illuminated reflection stage with the STAGE-RTL-T adjustable reflection and transmission stage. The Vivo provided bright, diffuse illumination with four tungsten halogen bulbs illuminating the sample. A 2 meter VIS-NIR fiber with a 600 micron core diameter (QP600-2-VIS-NIR fiber) arranged at a 45 degree angle relative to the tungsten halogen bulbs was used for the measurement of diffuse reflection from the fruit. Reference measurements were made with a WS-1 diffuse reflection standard. The dark measurement was made with the lamps turned on and an empty optical stage. The stage was shielded from overhead illumination during the dark measurement with a black shroud. The setup used for the measurements is shown in Figure 1.
NIR diffuse reflection was measured for whole fruit samples with measurements made at four different locations on the fruit. The fruit was placed on the magnetic ring of the Vivo optical stage to keep the fruit from rolling off the stage. Multiple measurements were made due to the variable nature of fruit — bruising, non-uniformity in color and differences in sugar content (due to differences in sun exposure) all lead to NIR spectral differences. To account for the inhomogeneity and variations in the fruit surface, many more measurements should be made at different points on the fruit surface.
NIR diffuse reflection measurements were made for whole ripe and unripe mangoes and avocados. In Figure 2, the average of the spectra measured at four locations on each piece of fruit is shown for two mangoes and two avocados. Multiple spectra (n=4) were recorded for each piece of fruit to account for the inhomogeneity of the fruit. These spectra demonstrate that even spectra for the same fruit type show variability across the spectral region with the avocado more consistent in the region above ~1100 nm. While the spectral features are similar for both types of fruit, differences in magnitude are observed throughout the spectra. Sampling additional locations on the surface of the fruit would help to average out variability for a given piece of fruit and improve the accuracy and repeatability of the results. Fortunately, the speed of the NIR technique provides the option to sample a larger surface area of the fruit with multiple measurements without the need for lengthy measurement times.
Notably, spectral features observed in these diffuse reflection spectra arise from a combination of phenomena depending on the amount of light scattered from the surface of the fruit and the penetration depth for NIR light into the sample. Light that is not scattered by the surface of the fruit passes through the peel and enters the fruit where it can be absorbed based on chemical composition. While diffuse reflection measurements are relatively straightforward to make, diffuse reflection from a variable rounded sample like a piece of fruit results in a complicated spectrum requiring carefully constructed interpretation models to extract quantitative information.
The spectra for peeled versus unpeeled ripe avocados and mangoes are shown in Figures 3 and 4. In the case of the avocado shown in Figure 3, the spectral features are more pronounced for the peeled avocado than the unpeeled avocado. This may result from less reflection of light by the peel, which increases absorption based on the chemical composition of the avocado.
For the mango shown in Figure 4, the effect of peeling the fruit is similar to the spectral differences observed for peeled and unpeeled avocados. But the impact of removing the peel is not as significant for the mangoes (there is less smoothing due to light reflected from the peel). The spectral differences observed when an avocado or mango is peeled suggest that different fruit peels have different properties that impact the overall fruit spectrum either through chemical composition or reflection properties.
The spectra for ripe versus unripe avocados and mangoes are shown in Figures 5 and 6. In Figure 5, the unripe avocado spectra are very consistent in the region from ~1900-2500 nm. The spectrum for the ripe avocado in this region flattens relative to the unripe avocado.
In Figure 6, the ripe versus unripe mango spectra are very similar with only very slight differences occurring between ~1900-2500 nm. These differences are most likely related to differences in sugar and starch content as the fruit matures. While many of the spectral changes are very subtle, a carefully constructed calibration model and a good sampling approach could be used to extract more quantitative information on fruit maturity from these spectra.
NIR spectroscopy is a powerful measurement tool for the characterization of agricultural samples. In the case of fruit, long NIR wavelengths where absorption is weak allow sampling through the peel of the fruit. This also makes sample preparation unnecessary. Combine these advantages with the ability to make rapid measurements and NIR spectroscopy becomes a great option for online measurement of fruit.
While the spectral data shown here illustrate the qualitative differences between avocados and mangoes at different stages of maturity, more quantitative information on fruit quality could be extracted from these spectra using an appropriate chemometric model and careful sampling to account for the inhomogeneity of the fruit.
Near-infrared Spectroscopy in Food Analysis, Brian G. Osborne, Encyclopedia of Analytical Chemistry, 1986