Professional Documents
Culture Documents
a r t i c l e i n f o a b s t r a c t
Article history: Rapid development in apple wine industry has raised a growing need for effective fermentation process
Received 25 May 2015 monitoring. The alcohol strength and titratable acidity have been used as indicators for process control
Received in revised form and optimization in apple wine making. The objective of this study was to investigate the efcacy of
29 September 2015
using near-infrared (NIR) spectroscopy to monitor and assess the process parameters during apple wine
Accepted 5 October 2015
Available online 9 October 2015
fermentation. A simple and rapid NIR spectroscopy method was developed for simultaneous determi-
nation of alcohol strength and titratable acidity in apple wine. The selected spectral regions of 6101.9
e5446.2 cm1, and 11,995.4e7498.1 cm1 were pretreated by second derivative (SD) and straight line
Keywords:
Near-infrared spectroscopy
subtraction (SLS) for alcohol strength and titratable acidity, respectively, prior to developing calibration
Apple wine models using partial least squares (PLS) regression with cross-validation. The highest R2c and the lowest
Monitoring RMSECV in the calibration set were obtained for the alcohol strength (0.923 and 4.63 mL/L) and for the
Fermentation titratable acidity (0.930 and 0.264 g/L). The NIR calibration models showed good correlation of deter-
Process control mination and low predictive errors. Application of the NIR calibration models demonstrated the feasi-
bility of NIR spectroscopy to be used as a quality control tool for monitoring the apple wine fermentation
process.
2015 Elsevier Ltd. All rights reserved.
1. Introduction 2003; Ye, Yue, Yuan, & Li, 2014; Zhang, Fang, & Li, 2011), but
these methods generally had the disadvantages of sample prepa-
Apple wine is widely used as a fermented alcoholic beverage in ration, reagent consumption and complicated operation with time-
most countries. The steadily-growing diversity of commercial apple consuming that might hamper the quality of nal apple wine. With
wine, coupled with increasing consumer demand, has prompted a the development of brewing technology and the demand of quality
need to monitor their major chemical composition changes for control, it is important to perform fast and accurate determination
process control and quality assessment in order to better manage of target composition for cost saving and online monitoring prod-
fermentation process. Alcohol strength and titratable acidity have uct quality (Blanco, Peinado, & Mas, 2004).
been used as indicators during apple wine fermentation process for Nowadays, apple wine brewing industry needs to introduce and
quality control and optimization in apple wine industry; hence, it is develop more advanced analysis method online in order to get
of great importance to monitor the changes of alcohol strength and reliable feedback data in time, which applied to guide the
titratable acidity in fermentation process for improving apple wine fermentation process. The monitoring of a fermentation process
quality. A variety of methods have been developed for monitoring often requires fast and frequent measurements, which makes Near-
the parameters of alcohol strength and titratable acidity in previous infrared (NIR) spectroscopy an excellent candidate for online
research, including chemical assays, pH meters, and high pressure analysis to provide rapid determination and straightforward
liquid chromatography, etc. (S.A.C., 2008; Satora, Tarko, Sroka, & monitoring. Bioprocess monitoring is becoming one of the most
Blaszczyk, 2014; Wilson, Maguer, Duitschaever, Buteau, & Allen, challenging applications of NIR spectroscopy (Lachenmeier, 2007).
With the application of chemometric techniques, a single spectrum
can be subjected to many different calibration models, to measure
* Corresponding author. many compositions in various types of food (Shen et al., 2010). NIR
E-mail address: bzpeng@mail.hzau.edu.cn (B. Peng). spectroscopy is a widely used rapid and non-destructive technique
http://dx.doi.org/10.1016/j.lwt.2015.10.018
0023-6438/ 2015 Elsevier Ltd. All rights reserved.
B. Peng et al. / LWT - Food Science and Technology 66 (2016) 86e92 87
for quality measurements and control in food and agricultural about 30 mL of samples were discharged out directly from fer-
product analyses (Antonucci et al., 2011; Chen, Ding, Cai, Sun, & mented supernatant. One hundred and sixty samples were
Zhao, 2012; Chen, Zhao, Fang, & Wang, 2007; Grassi, Amigo, collected from eight trials for developing the models of quantifying
Lyndgaard, Foschino, & Casiraghi, 2014; Pouliot, Paquin, Martel, the alcoholic strength and titratable acidity, while another trial
Gauthier, & Pouliot, 1997; Xie, Ye, Liu, & Ying, 2011), and is with twenty collected samples was used to evaluate the feasibility
currently an ideal alternative to traditional analytical techniques for of FT-NIR spectroscopy in monitoring apple wine fermentation
monitoring bioprocess (McLeod et al., 2009). With the application process.
of chemometric techniques, a single spectrum can be subjected to
many different calibration models, to measure many compositions 2.2. Chemical analysis
(McLeod et al., 2009), and this modern analytical technology has
been used as a well-suited method for general process real-time The performance of a quantitative NIR model is mainly depen-
monitoring, which is of great interest for many wine makers to dent on the performance of reference method (Bock & Connelly,
conduct process control. Di Egidio, Sinelli, Giovanelli, Moles, and 2008). Poor precision and accuracy of the reference method will
Casiraghi (2010) obtained good calibration models for the predic- limit the performance of the NIR model. Therefore, it is important
tion of the main compositional changes with FT-NIR to evaluate to ensure the targeted parameter as close to the true value as
alcoholic fermentation online in red wine. Buratti et al. (2011) possible. In this study, samples were centrifuged at 5000 rpm for
demonstrated that the non-destructive method of NIR are suit- 5 min and supernatants were collected for chemical analysis.
able for the monitoring of must-wine fermentation giving crucial Alcoholic strength was measured by using an PT-1 alcoholmeter
information about the quality of the nal product in agreement (Pute company, Beijing, China), immersed in a distillate at a tem-
with chemical parameters. Giovenzana, Beghi, and Guidetti (2014) perature of 20 C; titratable acidity was determined by acidebase
carried out a preliminary study to verify the possibility of titration method according to SAC standard GB/T 15038 (S.A.C.,
employing a device based on NIR spectroscopy, directly on the 2008) and the results were expressed as malic acid. All reagents
production line of craft beer. Grassi et al. (2014) investigated the used were of analytical grade, and related solutions were prepared
capability of FT-NIR spectroscopy to monitor and assess process by ultrapure water produced with a millipore purication system
parameters in beer fermentation with two different yeast strains at (Millipore, Mississauga, Canada) in the experiments. All analyses
different temperatures, and the results showed that FT-NIR spec- were done in triplicate.
troscopy demonstrated to be a perfectly suitable quantitative
method to be implemented in the production of beer. NIR spec- 2.3. Spectrum collection
troscopy has already shown promise as a rapid and non-destructive
method for determining various compounds simultaneously in the FT-NIR spectra collected directly from the supernatant of apple
fermentation industry (Liang et al., 2013). Nevertheless, less wine samples using a MPA FT-NIR spectrometer (Bruker Optics,
research exists on the application of NIR spectroscopy for moni- Germany) in the range of 12,000 to 4000 cm1, which equipped
toring and assessment of changes in relevant physico-chemical with an InGaAs detector and a liquid ber optic probe with a 2 mm
parameters in the apple wine fermentation process. Therefore, path length. An air background was employed as a probe reference,
much more intensive work is needed to really evaluate the feasi- and the liquid probe directly immersed into samples for collecting
bility of NIR spectroscopy in monitoring apple wine fermentation the spectra at 25 C. The spectral resolution was set at 8 cm1 with
process. The objectives of this study were to develop fourier an interval of 2 cm1 and the spectra were recorded performing 20
transform near infrared (FT-NIR) spectroscopy calibration models scans for both the reference and tested samples. The spectra were
for predicting alcohol strength and titratable acidity of apple wine recorded in triplicate and an average spectrum was collected to
in order to monitor fermentation process. In addition, the feasibility ensure an adequate signal-to-noise ratio for the subsequent anal-
of using FT-NIR spectroscopy to monitor these two indicators ysis. The NIR instrument control, spectra scanning, and initial data
simultaneously during the fermentation of apple wine was processing were performed by using OPUS software version 5.5
explored. (Bruker Optics, Germany).
2.1. Apple wine fermentation and sampling NIR spectra preprocessing in multivariate calibration is still trial
and error due to the lack of sufcient prior information and an
Ripe Fuji apples used in this study were harvested from different incomplete knowledge of the raw data (Xu et al., 2008). How to
commercial orchards located at Luochuan county in northwest select a proper preprocessing method depends largely on the
China, and stored in a cold room at 4 C until use. Apple juice was expertise and experience of practitioners. To establish best cali-
obtained by squeezing the mixed apples from different orchards bration model, spectra data from samples need to conduct different
from LZ juice extractor with a vacuum pressing and ltration sys- treatments for reducing noise interference and increasing the res-
tem in room temperature (Gongda company, Wenzhou, China), olution of small absorbance bands (Huang, Yu, Xu, & Ying, 2008).
during which the sodium bisulte of 100 mg/L was added to apple Chemometric analysis is usually used as a tool to correlate the
juice in order to inhibit the bacteria growth immediately (Peng, effective information in the NIR spectra signal to a number of latent
Yue, & Yuan, 2008). The total sugar and pH of all juice samples variables or factors, constrained by a set of calibration reference
for fermentation were then adjusted to 200 g/L with saccharose and data (Massart et al., 1997). Partial least squares (PLS) regression is a
3.4 with 0.5 M hydrochloric acid, respectively. A total of nine apple well-established chemometric technique whereby factors are
wine fermentation trials were conducted using three different derived by taking into account the variation in the spectral data
yeasts (Saccharomyces cerevisiae 1750, PF12 and WP45) at three that is relevant for explaining variation in the characteristics of
temperatures of 17, 20 and 23 C according to dry type brewing interest in the original samples (Williams, 2001).
technology in laboratory. The apple wine samples were collected In this study, spectra preprocessing was made by straight line
right after yeast inoculation (0 h, starting time), and then every 12 h subtraction (SLS), rst derivative (FD), second derivative (SD),
until the 10th day of fermentation in each trial. When sampling, vector normalization (VN), minemax normalization (MMN),
88 B. Peng et al. / LWT - Food Science and Technology 66 (2016) 86e92
Table 1
Analysis for chemical reference values of alcohol strength and titratable acidity in samples.
n: the number of samples; CV: coefcient of variation; PSE: pooled standard error.
multiplicative scattering correction (MSC), and the combination of 3. Results and discussion
every two methods using OPUS software (v.5.5 Bruker Optics,
Germany). The number of latent factors used in the calibration 3.1. Chemical analysis
models is of critical importance to avoid under or over tting of the
data and to ensure that the models are optimized for the intended Samples for establishing NIRS model should be representative of
purpose of the procedure (Inon, Llario, Garrigues, & de la Guardia, the commercial fermentation process of apple wine, and have the
2005; Valderrama, Braga, & Poppi, 2007). expected variability within the scope of the NIR calibration proce-
dure. The calibration and prediction sets contained 120 and 40
samples, respectively, which were selected by the KennardeStone
2.5. Establishment and evaluation of NIRS models algorithm in each set that covered the whole range of each chem-
ical parameter (Kennard & Stone, 1969; Valderrama et al., 2007).
For establishment of NIRS model, calibration is generally per- The samples of prediction set were selected as external validation
formed by developing a calibration model that relates target (not used in calibration procedures) to evaluate the prediction
composition concentration to absorbance spectra for a set of ability of calibration models.
reference samples in calibration set (Cozzolino, Parker, Dambergs, The range, mean, coefcient of variation (CV) and pooled stan-
Herderich, & Gishen, 2006). In this study, the calibration models dard error (PSE) for the alcohol strength and titratable acidity
of alcohol strength and titratable acidity were developed using PLS measured in both calibration and prediction sets were listed in
regressions with leave-one-out cross validation, which it makes a Table 1. A calibration set containing 120 samples was used for
subset of the samples leaving one out and trying to predict the one model development in the ranges from 0 to 126.3 mL/L for alcohol
that was left out (Norgaard et al., 2000). The performance of cali- strength, and from 2.08 to 8.92 g/L for titratable acidity, respec-
bration models was evaluated in terms of determination coefcient tively. The ranges in values measured were in accordance with
(R2) and root mean square error of cross-validation (RMSECV), typical values expected in apple wines (Satora et al., 2014; Ye et al.,
among which the R2 is a way of describing the strength of the 2014). Concentration spots of alcohol strength and titratable acidity
relationship between two variables, and the larger the R2 is, the in samples were plotted in Fig. 1. As can be seen from the Fig. 1,
more variability is explained by the calibration model; the RMSECV there is a positive relationship between alcohol strength and
is usually dened as the standard deviation of differences between titratable acidity. This is because both of the two substances are
spectral data and reference values in the cross-validation sample yeast metabolites in fermentation process, and can thus accumu-
set (Di Egidio, Olivieri, Woodcock, & Downey, 2011; Williams, late continuously in apple wine samples. The statistics data showed
2001). Generally, a better model should have higher R2 and a considerable degree of variations in the content of alcohol
smaller RMSECV. In addition, to evaluate the predictive capability of strength and titratable acidity, as indicated by the range and CV
the calibration model, root mean square error of prediction
(RMSEP), which indicates the total error in prediction, should be
acceptable when the model is applied to generate predictions
outside the calibration set. Moreover, the ratio of standard devia-
tion to the standard error of prediction (RPD) was also employed as
an indicator of NIRS predictions in this study. And higher RPD
values usually indicate better NIR predictions (Williams, 2001).
Generally, a good model should possess higher values of R2 and
RPD, and lower values of RMSEC and RMSEP (Bao et al., 2014). Only
optimal models are selected and discussed in this study.
The related equations dened as
0 1
B Pn
B yTi yPi 2 C
C
R2 B1 Pi1 C;
@ n
yTi ya 2 A
i1
s s
Pn 2 Pm 2
i1 yTi yPi i1 yTi yPi
RMSECV ; RMSEP
n m
where yTi is the reference value of the parameter for the i th sample
of the calibration set or prediction set; yPi is the predicted value
obtained by NIRS model; ya is the average value of reference; n, m Fig. 1. Concentration spots of alcohol strength and titratable acidity in samples for
are the number of samples in calibration set and prediction set, developing FT-NIRS models of alcohol strength and titratable acidity in order to
respectively. monitor apple wine fermentation process.
B. Peng et al. / LWT - Food Science and Technology 66 (2016) 86e92 89
Table 2
Statistics of calibration models of alcohol strength and titratable acidity in apple wine samples.
Parameter Spectral region Preprocessing method Rank Calibration set Prediction set
SD: second derivative; SLS: straight line subtraction; RMSECV: root mean square error of cross validation; R2c : determination coefcient for calibration set; R2p : determination
coefcient for prediction set; RPD: residual predictive deviation.
values in Table 1. The large CV values of alcohol strength (67.6%, best conditions including the spectral region, preprocessing
58.7%, respectively for calibration and prediction sets) and titrat- method and rank, were selected based on the smaller rank value
able acidity (32.7%, 24.3%, respectively for calibration and predic- and RMSECV for the calibration models according to PLS algorithm.
tion sets) were possibly related to different fermentation The statistics of optimal conditions are given in Table 2. The best
conditions being studied. Therefore, the variability of sample data spectral regions selected for developing calibration models of
in both calibration and prediction sets was considered suitable for alcohol strength and titratable acidity were 6101.9e5446.2 cm1,
developing stable and reliable NIR calibrations for alcohol strength 11,995.4e7498.1 cm1, respectively. The proles of selected spectral
and titratable acidity. regions preprocessed by second derivative (SD) and straight line
subtraction (SLS) for alcohol strength and titratable acidity,
respectively, are shown in Figs. 2 and 3. From Figs. 2 and 3, more
3.2. Selection and preprocessing of NIR spectra positive information and less noise were identied and extracted
from the differences of the spectra pretreated by SD or SLS, and thus
Preprocessing of raw near-infrared spectra (NIRS) is indispens- these pretreatments had contributed most to precision of each
able because the measured spectra are subject to signicant noises, model.
baselines and other irrelevant information, which may affect the
predictive ability of NIRS model (Blanco et al., 2004). Hence, it is
imperative to perform the spectral range selection and pretreat- 3.3. Calibration and validation of models
ment for visualizing and extracting relevant information from the
spectra best suited for calibration model. In this study, the auto- The calibration models of alcohol strength and titratable acidity
mation optimization function of the OPUS-QUANT2 software was were developed using PLS regressions with leave-one-out cross
used to optimize the calibration conditions for developing best validation. The PLS regression is a technique that constructs new
calibration models of alcohol strength and titratable acidity. The predictor factors derived by taking into account the variation in
Fig. 2. The prole of selected spectral regions of 6101.9 ~ 5446.2 cm1 before and after pretreatment by second derivative (SD) for developing calibration model of alcohol strength
in apple wine fermentation process (a: the original selected spectra; b: the spectra after pretreatment by SD).
90 B. Peng et al. / LWT - Food Science and Technology 66 (2016) 86e92
Fig. 3. The prole of selected spectral regions of 11995.4~7498.1 cm1 before and after pretreatment by straight line subtraction (SLS) for developing calibration model of titratable
acidity in apple wine fermentation process (a: the original selected spectra; b: the spectra after pretreatment by SLS).
spectral data that is relevant for explaining variation in the char- 2005). The lowest RMSECV and the highest R2c in the calibration
acteristics of interest in original samples (Li, Zhang, & Atungulu, set were obtained for alcohol strength (4.63 mL/L and 0.923) with
2014; Williams, 2001), and it is also one of the standard methods removal of 5 outliers, and titratable acidity (0.264 mg/L and 0.930)
used to develop calibration equations to predict chemical compo- with removal of 6 outliers. Generally, values of R2 greater than 0.9
sitions of a sample from NIR spectra (Reeves, 2001). indicate excellent quantitative information for calibration model
Statistics for the calibration models developed for alcohol (Shenk & Westerhaus, 1996). To improve the stability of the models,
strength and titratable acidity in apple wines using FT-NIR spectra the rank must be appropriately chosen to avoid overt during
are shown in Table 2. Outliers in the calibration set were identied calibration. Fig. 4 shows the correlation between RMSECV and Rank
and excluded based on extreme leverages (Lillhonga & Geladi, in calibration by PLS regression with leave-one-out validation, and
Fig. 4. Correlation between RMSECV and Rank in calibration set by PLS regression with leave-one-out cross validation for developing FT-NIRS models (a: for alcohol strength; b: for
titratable acidity).
B. Peng et al. / LWT - Food Science and Technology 66 (2016) 86e92 91
Fig. 5. Correlation statistics between chemical reference value and FT-NIRS prediction value in calibration and prediction sets. The FT-NIRS models were developed by PLS
regression with leave-one-out cross validation using selected spectral regions of 11,995.4 ~ 7498.1 cm1 pretreated by SLS, and 6101.9 ~ 5446.2 cm1 pretreated by SD for alcohol
strength (a) and titratable acidity (b), respectively.
the optimum ranks are both 6 for alcohol strength and titratable suggest that the calibration models have good predictive capacity
acidity calibration models. The correlation plots between chemical for alcohol strength and titratable acidity during apple wine
reference value and FT-NIR prediction value for the two parameters fermentation, and this rapid NIR method could be taken to monitor
of apple wine in calibration set are shown in Fig. 5. The results show the apple wine fermentation process with sufcient feasibility.
that the NIRS calibration models have excellent precision. There-
fore, these two physicochemical parameters in apple wine might be
measured by FT-NIR spectroscopy. 4. Conclusion
In order to further evaluate the prediction capability of the
models, an external validation was conducted by applying FT-NIRS This study developed a simple and rapid NIRS method for
models to the 40 samples of prediction set. Table 2 shows that simultaneous determination of alcohol strength and titratable
adequate small RMSEP values were obtained for alcohol strength acidity content in apple wine. The NIRS models showed a good
and titratable acidity (4.25 mL/L and 0.213 mg/L), meanwhile the R2p agreement between calibration and predication data with adequate
values (0.993 and 0.973) are closed to 1. The rate of prediction to high correlation of determination of R2 and relatively low predic-
deviation (RPD) could be an indicator of model validation and a tive errors of RMSEP. The results demonstrated that NIR spectros-
higher RPD would be deemed better for prediction capability of copy can used as an alternative method to rapidly analyze alcohol
model. A good calibration model should have a RPD value higher strength and titratable acidity, with a very high applicability for
than 3.0 (Williams, 2001). A RPD value of 6.5e8.0 is generally monitoring apple wine fermentation process in order to improve
considered as good for quality control purpose (Botelho, Mendes, & the quality of apple wine.
Sena, 2013). As shown in Table 2, RPD for the calibration models of
alcohol strength and titratable acidity (7.12 and 6.86) were
considered satisfactory. Additionally, the correlation plots between
chemical reference value and FT-NIR prediction value for the two
parameters of apple wine in prediction set are shown in Fig. 5. The
results indicate that the calibration models are adequate as a tool to
determine the alcohol strength and titratable acidity with accurate
performance during apple wine fermentation. Similar results were
obtained by Ye et al. (2014) for the determination of total acidity
with the determination coefcients of 0.98 and the RMSEP of 0.021
in apple wine by FT-NIRS transmission mode.