8290614327

7. Opublikowane badania własne

y Oncl et al. / Talmta I3« (2015) 64-70    65

methods tKat are bascd on casy-to-perform on-łine measuremenis that have no requirements for a special laboratory prenearment wirh reagenrs are greatfy desired.

A good candidate is a method based on the multivariate calibration of IR spectra (non-selective signals with respect to the panku lar antioxidant compounds). Lu and Rasco (2012) reported on near- and mid-IR (12.800-400 cm“ *) spectroscopy as a suitable techniąue to evaluate the TAC of selected food commodities 112). That is why the IR measurements for vegetables 113|. frnits (141. beverages (151. etc. have been recently proposed as a replacement for convrntional assays. This approach is ctaimed to be accurate. non-destructive and to substanbally reduce che amount of preparative work that is required to analyze samples on one hand, while it is sensitwe to the presence of water in the samples on the other. Therefbre. the analysis of either extracts or yegetable and fruit pulps still presents a challenge. Moreover. the IR radiation is relatively Iow (16] for frnits or vegetables with a thick skin or occlusive flesh penetration.

Another promising methodołogy that uses non-sełective fluores-cence spectra and multivariatp modeling has been proposed to evaluate the TAC of coffee and peppermint extracts (17|. Each sam ple was excited using radiation at diffcrent energy levels (various excitation wavelengths) and the emission spectrum was collected for each excitation wavelength. The excitatlon-emission fluorescence matrices (EEMs) that were collected are examples of second order signals and their use can facilitate the calibration of the constituents of samples (or some specific features like color. ash content, and shelf life) when interfiering compounds are present in the sample marrix (18.19). The relatively Iow prediaion errors for the TAC of the multivariate calibration models that use EEMs (equal to 625% and 9.08% for coffee and peppermint extracts. respectively) indicate the potential of the proposed methodołogy. The preliminary studies were focused only on the water extracts of samples 117).

The aim of this study is to develop a suitable approach for the determination of the TAC levels of intaa food samples that woułd allow for the possibility of routine on-line testing. Tomato pastę, which is an integral part of the human diet worłdwide. was chosen as the subject of this study. For the collecrion of intact samples. EEMs were registered using a fi ber optic probe in a selected UV-vis rangę. In order to compare the results that were obtained from EEMs with those that were obtained from the methodołogy proposed in the literaturę for the IR measurements, the mid-IR spectra (4000-400cm ') for the same collection of samples were also registered Antioxidant water extraction conditions (time and temperaturę) were optimized using the central composite design (20) in order to guarantee the optimal TAC determination. The TAC parameter in this study was evaluated using the oxygen radical absorbance capacity assay. the ORAĆ assay (21). Even though. there are no strict criteria for the selection of an assay. the ORAĆ assay seems to be selected the most among the assays (22.23). It should be emphasized the TAC of water extracts discusscd in this study refer to the total content of hydrophilic antioxidants extracted Prom selected samples.

The total phenolic content (TPC) is another parameter that refers to the antkaidant content in samples. In this study. the TPC was determined using the Fołin-Gocalteu reagent (24). Partia! least squares regression. PLSR 125] was used to model the TAC (or TPC) as a function of the spectra for respective intaa food samples In addidon. N-way partial least squares regression models (extension of PLS). N-PLS [ 26). were developed for the collection of the EEMs spectra 2. Experimental

2.f. Origi/i of the samples

The tomato pastes that are available on the Polish market were purchased in local Stores. They were produced by different producers that were located in different p rod uch on places (nine tomato pastes originated from Poland. three from Itaiy and one from Croaria). The packaging matetials that were used were also different ()ars. cans and cardboard boxes). A single sample was prepared using the water extraction of OS g tomato pastę under optimal extraction conditions. Each pastę was analyzed in ttiplicate resulong in a total of 39 samples.

22. Determining the optimal conditions for the extraction procedurę

To reduce the amount of hazardous solvents for the environ-ment. distilled water was used for the extractions. Optimal exrraction conditions were determined using the central composite design. CCD. with rwo faaors (time of sonicarion and temperaturę). Five levels were considered for each factor. The ranges and coded levels of investigated faaors are presented in Table 1.

A total of 20 experiments were canied out. In order to evaluate the pure experimental error ratę. experiments with nos. 1 -8 were repeated twke and the experiment at the central point was repeated four times. The ORAĆ parameter was the response variabłe, y. that was determined in each experiment. In generał, the following quadratk model was considered:

y = b₀+6,xi + bjx₂ + b>x? +b₄x§ + b₅x,x₂+e    (1)

Analysis of yariance. ANOVA, with the rype III sum of squares was adopted to determine the significance of each regTession coefficient of the ąuadratic equation. ANOVA was also used to check the adequacy of the finał model (the significance of lack-of-fit), sińce the es ti ma bon of the pure measurement error was available.

2.3. Evaluarion of the total antioxidant capacity using the ORAĆ assay

The oxygen radical absorbance capacity. the ORAĆ assay (21J. is measured as the damage to a fluorescence probe that is caused by the free radicals generated from AAPH. Reaaions for a sample with the analytes (antioxidants) and for a sample without analytes are monitored in parallel and the TAC is evaluated on the basis of the area between the two curves that are registered.

The ORAĆ assay was performed using a 96-well piąte. Fluoresce was measured at hxed excitation and emission wavelengths

Table 1

Desgn of ck pen mcm.

Run (rjodom	Time	Temperaturę Time todetf		Trapcntitr
oiitr)	(min)	CC)	(«.)	rodni (jr,)
6	5	34	-1	■
3	S	34	- 1	|
16	13	34	t 1	— 1
U	13	34	> 1	- |
7	5	72	- 1	♦ 1
17	5	72	- 1	♦ 1
14	13	72	+ 1	♦ 1
8	11	72	1 1	♦ 1
S	3	53	- 1.414	0
19	3	53	- 1.414	0
IS	IS	53	+ 1.414	0
1	15	53	> 1.414	0
2	9	26	0	- 1.414
4	9	26	0	-1.414
18	9	80	0	1.414
13	9	80	0	1.414
9	9	53	0	0
20	9	53	0	0
10	9	S3	0	0
12	9	53	0	0

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