Anders Et Al 2015

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Scientia Horticulturae 195 (2015) 216–225

Contents lists available at ScienceDirect

Scientia Horticulturae

journal homepage: www.elsevier .com/ locate /scihor t i

Raspberry fruit quality changes during ripening and storage asassessed by colour, sensory evaluation and chemical analyses

Jon Anders Stavang a, Sabine Freitag b, Alexandre Foito b, Susan Verrallb, Ola M. Heidec,Derek Stewart b,d, Anita Sønsteby d,∗

a Department of Biology, University of Bergen, NO-5008 Bergen, Norwayb Environmental andBiochemical Sciences, The James Hutton Institute, Invergowrie, DundeeDD25DA, SCT, UK c Department of Ecology andNatural ResourceManagement, NorwegianUniversity of LifeSciences,NO-1432 Ås,Norwayd NIBIO, Norwegian Institute for Bioeconomy Research, NO-1431 Ås,Norway

a r t i c l e i n f o

Article history:Received 20 February 2015Received in revised form 26 August 2015Accepted 28 August 2015

Keywords:RaspberryColorSensory evaluationChemical compositionRipeningStorage

a b s t r a c t

In order to identify the optimal harvest time and monitor changes in raspberry (Rubus idaeus L. cv. GlenAmple) fruit quality during ripening and storage, quality was assessed and compared by physical, chem-ical and sensory fruit quality criteria. Visual classification of fruit colour according to the Natural ColourSystem (NCS) chart and by physical measurement of fruit adherence to the receptacle or fruit compres-sion resistance yielded parallel and highly significant results. The light red colour stage correspondingto NCS S code 3060-Y90R was identified as the optimal harvest stage for commercial fresh marketing of the ‘Glen Ample’ cultivar. Fruit harvested at this stage developed the same chemical and sensory quali-ties as in situ matured fruits and maintained high sensory quality after 8 days of storage in the dark at2–3 ◦C. As the fruits mature, the concentration of titratable acids decreases, whereas the concentrationsof anthocyanins and the sugar:acid ratio increase in parallel with colour development. While correlationanalysis revealed a correlation between sensory traits like sweetness and acidity with sucrose and thesugar:acid ratio, respectively, the overall fruit tastefulness was not strongly correlated with any specificphytochemical component, thus illustrating the complex nature of this sensory trait. Due to its ease of

performance, picking raspberry fruits related to a standardised colour chart is recommended for pickingraspberry fruits with optimal quality.

© 2015 Elsevier B.V. All rights reserved.

1. Introduction

Redraspberry(Rubusidaeus L.)isgrownthroughoutthetemper-ate regions of the world as a commercially important berry crop.The fruit is highly valued for its flavour and high content of poten-tially importanthealth-beneficial constituents (Mullen et al., 2002;Liu et al., 2002; Anttonen and Karjalainen, 2005; Rao and Snyder,2010). It is a perishable commodity, and although new cultivarswith firmer fruit have been released (Finn et al., 2008) the shelf life

of raspberry is generally short. Identification of the optimal matu-rity stage for harvesting and correct post-harvest handling andstorage of the fruit, are therefore,essential for successful marketingof fresh consumption raspberries.

Fruit colour and adhesion to the receptacle plug are the maincriteria used by the producer for practical assessment of the rightmaturitystageforharvesting,whilecolourisalsothemaincriterion

∗ Corresponding author at:E-mail address: [email protected] (A. Sønsteby).

used by the consumer to judge fruit quality. The main contribu-tors to the red colour of the raspberry fruit are the anthocyanins.Only cyanidin-and pelargonidin-type anthocyanins are present inred raspberry, the former type predominating (Wang et al., 2009;Remberg et al., 2010; Mazur et al., 2014). Their synthesis is influ-enced by a number of environmental factors in both green leavesand in fruits. According to Grisebach (1982), light is the mostimportant factor influencing anthocyanin biosynthesis in plantsin general, and in vegetative tissues, anthocyanin biosynthesis is

induced by UV light as an important photo-protective mechanism(Steyn et al., 2002). In ‘Glen Ample’ raspberry fruit, the concentra-tion of total monomeric anthocyanins (TMA) was not significantlyaffected by post-flowering growth temperature in the 12–24◦Crangeorbyphotoperiodat18 ◦C (Remberg et al., 2010; Mazur et al.,2014). It is well known anddocumented however, that postharvestcolourchangesandanthocyaninsynthesistakeplaceinimmaturelyharvested fruit of red raspberry (Wang et al., 2009) as well as inother berry species (Kalt et al., 1993; Sachs and Shaw, 1993), thusdemonstrating de novo synthesis in detached fruits. Synthesis canoccur in darkness, but the rate is slightly enhanced in light (Austin

http://dx.doi.org/10.1016/j.scienta.2015.08.0450304-4238/© 2015 Elsevier B.V. All rights reserved.

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J.A. Stavang et al. / Scientia Horticulturae 195 (2015) 216–225 217

et al., 1960; Wang et al., 2009). While raspberry fruit anthocyaninconcentration increases throughout ripening, the total phenolicconcentration was found to decrease from the green to the light-red (‘pink’) stage, and thereafter to increase again until maturity(Wang andLiu, 2000). However, because fruit weight increases sig-nificantly during ripening, mainly due to increased water content,a significant dilution of all soluble fruit constituents is also takingplace during fruit ripening. This is confounding much of the realconcentration changes in fruit constituents that are taking place.As pointed out by Remberg et al. (2010), this bias is corrected forwhen data are expressed on dry weight basis.

While a number of investigations have examined the effects of fresh and frozen storage on raspberry fruit chemical composition(e.g. Kalt et al., 1999; Mullen et al., 2002 and references therein),experimental attempts to define and describe maturity stages asguidelines for selection of the best harvest time for raspberriesare rare. Some studies in raspberries (Krüger et al., 2003; Krügeret al., 2011) have been restricted to the analysis of three stages of development: semi-ripe, ripe and over-ripe. Krüger et al. (2003)observed in four cultivars that the ripening stage had a large effectonfirmness,titratableacidity(TA)andfruitcolour,whereasstorageconditions during three days had effects on the content of solublesolids (SS) and suitability for shipping. A subsequentchemical anal-ysis of redraspberry (cv. ‘Tulameen’) at the three different stages of development, revealed that acidity decreased significantly, whiletotal anthocyanins increased significantly with maturation stagewith cyanidin-3-sophoroside, cyanidin 3-rutinoside and cyanidin3-O-glucoside levels increasing with maturation but not cyanidin3-glucosylrutinoside (Krüger et al., 2011). Further sensory analysisrevealed that ripening had an effect on odour and taste whereasthe effects of storage were small in comparison (Krüger et al.,2003). Hence, it was suggested that semi-ripe fruits could haveimproved suitability for shipping and some sensory assessments(Krüger et al., 2003).

In order to determine howearly fruits can be harvested andstilldevelop acceptable quality, Wang et al. (2009) harvested fruit of ‘Caroline’raspberries at fivematuritystages, arbitrarily classifiedas

5%, 20%, 50%, 80%, and100%maturity, andstudiedtheirfruitqualitychemical changes during storage in light and darkness at 24/16◦C(day/night) temperature. The authors found that fruit harvested at5% or 20% maturity never developed the levels of SS, TA and sug-ars found in ripe berries at harvest, while those harvested at 50%or 80% maturity attained qualities comparable to in situ maturedberries. Storage in light enhanced sugar and reduced acid contentsomewhat of 5% and 20% mature berries, but had negligible effecton fruit of more advanced maturity Theauthors conclude that rasp-berry fruit could be harvested as early as 50% maturity, when thefruits are firmer and less susceptible to mechanical injury, and stilldevelop a quality comparable to fully mature fruit. However, nosensory assessment of fruit quality was provided.

In an attempt to describe more precisely the fruit quality

changes taking place during raspberry fruit maturation and postharvest storage, andin particular, to define the right maturity stagefor harvesting, we have assessed and compared physical, chemicaland sensory fruit quality criteria that can be used for this purpose.It is hoped that this knowledge may benefit the fruit industry andthe producers as well as the consumers.

2. Material and methods

2.1. Plant material and cultivation

Long canes of the raspberry cultivar ‘Glen Ample’, producedas described by Sønsteby et al. (2009), were cropped in a green-

house at a commercial grower’s nursery located on the west coast

Fig. 1. The numbered colours selected for classification of the maturity stage atharvest in ‘Glen Ample’ raspberry.

of Norway during the spring 2011 (Frekhaug, 60◦31 N; 5◦14 E).The plants were tipped at a height of 180cm and grown in 3.5L pots in rows with 5 plants per running m. The distance betweenthe rows was 2.20 m. The plants were fertilized with a completenutrient solution containing a 2:3 mixture of CalcinitTM (15.5% N,19% Ca) and SuperbaTM Red (7–4–22% NPK+ micronutrients) (Yara

International, Oslo, Norway). The media electric conductivity (EC)was 1.3–1.6mS cm−1. The production started in midFebruary 2011andthe first berries were harvestedthe first week of May2011. Theberries used in this experiment were harvested on 17 May 2011.

2.2. Physical characterization of maturing raspberries

In order to pick berries with different degrees of maturity,five berry colours along an assumed maturity gradient were iden-tified and used as a reference during the picking. The coloursranged from orange/red (colour one), light red (colour two), red(colour three), dark red (colour four) and dark red/lilac (colourfive). These colours were visually classified according to the natu-ral colour system®Ó (NCS; http://www.ncscolour.com/en/natural-

colour-system/; see Fig. 1). Raspberries of the respective colourclasses were handpicked, 2–3 berries from each plant, from tworows in thegreenhouse,and placedin black,capped plastic contain-ers(200g). When full, thecontainer with berries was transported toa cold room (2–3◦C) within 30min. Four containers were collectedfor each colour class and used in the further analysis.

For further characterization of berries with different colours, wealso measuredthe force neededto removethe berryfromthe recep-tacle in relation to colour. To measure the pull force, wefirst placeda piece of 15cm duct tape around an individual berry so that theends met surrounding the edge of the berry and the tape stickingto the side walls of the fruit. The loop of the tape was open to allowconnection to the hook of a Pesola precision spring scale (PesolaAG,Baar, Switzerland; “Medio Line 300 g” scale for colour class 1–3

and” Light Line 100 g” scale for colour class 4 and 5). With the ducttape connected to the berry, it was slowly pulled off the recepta-cle. Maximum force monitored during the pull was recorded andthis was replicated 12 times for each colour class. We also recordedberry weight in relation to colour by weighing three replicates of 20 berries each on a technical balance (±0.1g).

In order to measure fruit firmness at harvest, we placed 50raspberries from each maturation stage into a 1-litre cylinder andapplied a load of 500g with an adaptor fitting exactly the cylin-der diameter on top of the berries. The volume of the berries wasrecorded before the load was added and after 6 min. From the vol-ume change, %-compression was calculated. These measurementswere done within 30min after harvest. After 8 d of storage thiscompression method was not applicable as the berries became too

soft and the compression method resulted in a fruit pulp.

http://www.ncscolour.com/en/natural-colour-system/;http://www.ncscolour.com/en/natural-colour-system/;http://www.ncscolour.com/en/natural-colour-system/;http://www.ncscolour.com/en/natural-colour-system/;http://www.ncscolour.com/en/natural-colour-system/;http://www.ncscolour.com/en/natural-colour-system/;http://www.ncscolour.com/en/natural-colour-system/;http://www.ncscolour.com/en/natural-colour-system/;http://www.ncscolour.com/en/natural-colour-system/;http://www.ncscolour.com/en/natural-colour-system/;http://www.ncscolour.com/en/natural-colour-system/;

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At the end of the day, the cooled berries were transported bycar in electric portable coolers to a cold storage room at the Bio-forsk Ullensvang research station. Here, berries were stored for 1d or 8 d in the dark at 2–3 ◦C until used for sensory evaluation, orfrozen at −40 ◦C for subsequent chemical analysis. To prepare theberries fortargetedchemicalanalyses, a pooledsample of 55–195gof berries from each colour class was freeze dried, ground up andhomogenized in a mortar, and shipped in vacuum sealed bags to

James Hutton Institute, Scotland.

2.3. Sensory evaluation

At day 1 and 8, berries were put into electric portable coolersand transported by car to the Oslo head office of BAMA, Norway’slargest private distributor of fruit and vegetables. There, staff of theadministration made up a sensory team with 9 members. The sen-sory panel followed a standard protocol routinely used in BAMAssensory evaluation of fruits. First,the berries were allowed to reachroom temperature. Then the panel gathered in a meeting roomand the judges of the panel individually evaluated taste of acidity,sweetness,freshness,and bitterness, overall tastefulness andvisualattractiveness of the berries, and gave the berries scores from 1 to

9 for each character. Each judge wrote their evaluation on a paperthat was collected for data analysis after the end of the evaluation.A score of 9 for instance for acidity, implied a very acid berry, anda score of 1 would imply that the berries were not acid at all.

2.4. Phytochemical content

2.4.1. Soluble solids and titratable acidityAt Bioforsk Ullensvang, raspberry juice prepared from frozen

raspberries from 2 replicates of 10 homogenized and filtered rasp-berries of each colour class, was measured for% SS by a tablerefractometer (Atago DBX-50; Atago, Tokyo, Japan). An aliquot(5mL) of this raspberryjuice was further diluted with distilled H2O1:6 and assessed for TA with a Titramaster 85 (Radiometer analyt-

ical, Villeurbanne Cedex, France). Results for TA are expressed as%citric acid equivalents.

2.4.2. Individual sugar, organic acid and polyphenolquantification

Allchemicals used foranalysiswere of HPLC grade: Acetonitrile,methanol (HIPerSOLV, Chromanorm, VWR, UK), acetic acid (Chro-manorm, Prolabo, UK), formic acid (Sigma Aldrich, Dorset UK) andultrapure water (Elga, UK). Polyphenolic standards for compoundsfound in raspberry extracts were purchased from ExtrasyntheseLtd. (Genay, France) including cyanidin-3-O-glucoside chloride,cyandidin-3-O-rutinoside chloride, cyanidin-3-O-sambubiosidechloride, cyanidin-3-O-sophoroside chloride, cyanin chloride,quercetin-3-O-glucuronide and hyperoside (quercetin-3-O-

galactoside). Morin, sugars (glucose, fructose and sucrose) andorganic acid standards (quinic acid, oxalate, succinic acid and citricacid) were all purchased from Sigma–Aldrich (Dorset, UK). Allchemical analysis data were expressed on dry weight basis.

2.4.2.1. Extraction and quantification of individual sugars andorganic acids. Freeze-dried material was weighed (60±2mg) intoa 2 mL Eppendorf vial and extracted at room temperature for30min as described by Mazur et al. (2014). In summary, sam-ples were extracted in 1.5mL of extraction solvent (50:49:1methanol:dionised water:formic acid) at room temperature for30min. After centrifugation 1 mL of the supernatant was heatedat80 ◦C for 10min. After a centrifugation step, 500L of the super-natantwasevaporated andresuspended in 1 mL ofdeionisedwater.

Organicacids(citrateandmalate)werequantifiedfollowingextract

dilution (1:20) by anion exchange HPLC on a Dionex IonPac AS11-HC 4×250 mm column [Dionex (UK) Ltd. Camberley, UK] fittedwith a 4×50mm guard column utilising gradient of NaOH in 10%methanol (see supplementary Table 1) as described in Mazur et al.(2014). Sugars were quantified following extract dilution (1:500)by anion exchange chromatography on a Dionex Carbopac PA-100250×4 mm column [Dionex (UK) Ltd. Camberley, UK] utilising anisocratic elution with 200 mM NaOH prepared in degassed waterataflowrateof1mL min−1 for 15min, as described by Mazur et al.(2014).

2.4.2.2. Extraction and quantification of individual polyphenols. Forextraction and quantification of individual polyphenols 100±2 mgof freeze dried berry powder was weighted into 15mL amber glassvials sealed with a screw cap containing a PTFE liner (SULPELCO,Sigma–Aldrich, UK) and extracted with 3 mL of a water, acetoni-trile and acetic acid mixture in the ratio of 60:40:1 at 20◦C for1-h in a rotary shaker as described by Mazur et al. (2014). Thesupernatantwas dilutedin1:10ratioutilisingextractionsolvedandsubsequently transferred into filter vials and sealed with a 45mPTFE line screwcap (Thomson Instrument Company, London, UK).

The chemical analysis of the berry extracts was performed ona high performance liquid chromatography (HPLC) system consist-

ing of a quaternary pump (Agilent 1260), a DAD (Agilent 1260), acolumntemperaturecontroldevice(Agilent1260)andanautosam-pler Thermostat (Agilent 1290) coupled to a Triple QuadrupoleMass Spectrometer (Agilent Technologies, Santa Clara, CA, USA)with the instrument settings described in Mazur et al. (2014).The chromatography was performed on a Phenomenex C18(2)2×150mm (4m) column fitted with a C18 4×2 mm SecurityGuardTM cartridge (Phenomenex, Torrance, CA, USA) at flow rateof 0.3mL min−1 utilising a gradient consisting of 3 phases (seesupplemental table X). Individual polyphenols were quantified byreference to an external calibration curve generated using pur-chased standards.

2.5. Data analysis

Using GenStat for Windows, 16th Edition [16.2.11713 (64-bitedition) VSN International Ltd., Hemel Hempstead, UK], an analy-sis of variance with linear model was applied to the data set whichtested for a linear relationship to colour. For the sensory, sugar,organic acid and polyphenol data a two-factor model was usedto test for significant differences between the storage treatment,colour classand the interaction. For adhesion,weight and compres-sion data a single factor model was applied to test for significantdifference in the colour variable.

For the sensory analysis the judges were treated as a randomeffect, and all the data for colour 5 was excluded in the statisticalanalysis of the sensory evaluations of acidity, sweetness,bitterness,freshness and overall tastefulness as the data for this colour was

missing for berries stored for 8 d.Pearson’s correlation analysis and regression analysis werestatistical methods of choice for the analysis of phytochemicalproperties. Results for regression analysis gave the following infor-mation: (a) whether the slope for regression on the colour wassignificantly different to zero when combining the two treatments(e.g. Colour at harvest and Storage) (b) whether the intercept of 1or 8 d storage for individual compounds was significantly differ-ent, while having the same slope and (c) whether slopes for shortand long term storage for individual compounds were significantlydifferent from each other. Furthermore, correlation analysis wasapplied in order to investigate the internal relationship betweenorganic acids, sugars and flavonoids. Finally, correlation analysiswas applied in order to analyse the relationship between sensory

scores and fruit chemical content.

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Table 1

Physical traits of ‘Glen Ample’raspberryfruits harvestedat fivedefined colour stages. Results from theANOVA with linearmodelare presented, firstly theANOVA test, thenthe test forlinearity given as the significance of the slope in relation to colour. Different letters indicate a significant difference.

Colouratharvest

Pull force(g)±SD(n =12)

Berry weight(g FW)±SD(n=3, 10in each)

Compression (%)±SD(3) (n= 3)

1 183.4±35a 4.9±0.30a 30.74a*

2 138.5±44b 5.3±0.06ab 28.30±4.3a3 94.7±35c 5.6±0.03b 32.70±1.7ab

4 53.7±16d 6.5±0.30c 34.69±1.1ab5 33.2±15d 6.7±0.50c 38.84±4.3b

Source of variation (ANOVA)P - Value colour

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Table 3

Means±SD of SS, TA and SS/TA ratio after one or eight days of storage of ‘GlenAmple’ raspberry fruits harvested at five defined colour stages. ANOVA results arepresented with P - value and s.e.d. of each model factor.

Storage Colour at harvest SS (%)±SD TA (%)±SD SS/TA±SD

1d

1 9.25±0.1 2.8±0.06 3.3± 0.052 9.95±0.1 2.7±0.04 3.7±0.093 9.38±0.2 2.4±0.05 4.0±0.164 10.00±0.0 2.1±0.06 4.7±0.14

5 9.70±0.0 1.9±0.01 5.2± 0.02Mean 9.66 2.4 4.28d

1 9.60±0.0 2.7±0.02 3.5± 0.032 9.45±0.1 2.4±0.01 3.9±0.023 10.10±0.0 2.1±0.04 4.8± 0.104 9.80±0.0 1.8±0.01 5.4± 0.025 9.00±0.0 1.6±0.05 5.5± 0.17Mean 9.59 2.1 4.6

Source of variation (ANOVA)P - Value, Storage (S) 0.058

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Table 5

Probability levels of significance (P - value) and standard errors (s.e.) arepresented forregression analyses of organic acids, sugars, anthocyanins and flavonols in raspberrysamples harvested at different colour (maturity)stages and stored for 1 d or8 d in darkness at 2–3 ◦ C.

Phytochemicals Colour at harvest Storage Colour at harvest×Storage

Malate 0.008 0.008

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A 1 d storage

C i t r a t e

M a l a t e

O x a l a t e

Q u i n a t e

F r u c t o s e

G l u c o s e

S u c r o s e

C y a n i n

C y - 3 - O - S o p h

C y - 3 - O - S a m b

C y - G l u c - R u t

C y - 3 - O - G l u c

P e l - 3 - O - S o p h

C y - X y l - R u t

P e l - G l u c - R u t

C y - 3 - O - R u t

H y p e r o s i d e

Q u e r c G l u c

Citrate 0.91 0.90 -0.38 0.35 -0.53 -0.78 -0.75 -0.83 -0.84 -0.89 -0.83 -0.87 -0.87 -0.86 -0.85 -0.83 -0.81

Malate 0.91 0.95 -0.47 0.47 -0.51 -0.83 -0.86 -0.94 -0.91 -0.90 -0.81 -0.83 -0.89 -0.77 -0.76 -0.67 -0.69

Oxalate 0.90 0.95 -0.21 0.47 -0.40 -0.64 -0.67 -0.80 -0.76 -0.76 -0.65 -0.67 -0.73 -0.62 -0.61 -0.66 -0.76Quinate -0.38 -0 .47 -0.21 -0.58 0.04 0.86 0.59 0.54 0.57 0.54 0.51 0.57 0.56 0.54 0.52 0.04 -0.14

Fructose 0.35 0.47 0.47 -0.58 0.50 -0.55 -0.14 -0.25 -0.20 -0.17 -0.01 -0.10 -0.15 -0.05 -0.01 0.24 0.19

Glucose -0 .53 -0 .51 -0.40 0.04 0.50 0.36 0.76 0.70 0.74 0.76 0.83 0.76 0.76 0.75 0.78 0.85 0.78

Sucrose -0.78 -0.83 -0.64 0.86 -0.55 0.36 0.84 0.85 0.87 0.86 0.81 0.86 0.87 0.83 0.81 0.48 0.36

Cyanin -0.75 -0.86 -0.67 0.59 -0.14 0.76 0.84 0.98 0.99 0.96 0.95 0.93 0.97 0.88 0.89 0.70 0.61

Cy-3-O-Soph -0.83 -0.94 -0.80 0.54 -0.25 0.70 0.85 0.98 0.99 0.97 0.92 0.92 0.97 0.85 0.86 0.71 0.68

Cy-3-O-Samb -0.84 -0.91 -0.76 0.57 -0.20 0.74 0.87 0.99 0.99 0.99 0.96 0.96 0.99 0.91 0.91 0.75 0.68

Cy -Gluc- Rut -0.89 -0.90 -0.76 0.54 -0.17 0.76 0.86 0.96 0 .97 0.99 0.98 0.99 1 .00 0 .96 0 .96 0.82 0.74

Cy-3-O-Gluc -0.83 -0.81 -0.65 0.51 -0.01 0.83 0.81 0.95 0 .92 0.96 0 .98 0.99 0 .99 0 .98 0 .99 0.86 0.74

Pel-3-O- Soph -0.87 -0.83 -0.67 0.57 -0.10 0.76 0.86 0.93 0 .92 0.96 0 .99 0 .99 0.99 0 .99 0 .99 0.83 0.71

Cy-Xyl-Rut -0.87 -0.89 -0.73 0.56 -0.15 0.76 0.87 0.97 0.97 0.99 1.00 0.99 0.99 0.96 0.96 0.81 0.72

Pel-Gluc-Rut -0.86 -0.77 -0.62 0.54 -0.05 0.75 0.83 0.88 0.85 0.91 0.96 0.98 0.99 0.96 1.00 0.86 0.72

Cy-3-O-Rut -0.85 -0.76 -0.61 0.52 -0.01 0.78 0.81 0.89 0.86 0.91 0.96 0.99 0.99 0.96 1.00 0.87 0.74

Hyperoside -0.83 -0.67 -0.66 0.04 0.24 0.85 0.48 0.70 0.71 0.75 0.82 0.86 0.83 0.81 0.86 0.87 0.96

Querc Gluc

-0.81

-0.69

-0.76

-0.14

0.19

0.78 0.36

0.61

0.68 0.68

0.74

0.74

0.71

0.72

0.72

0.74 0.96

B 8 d storage

C i t r a t e

M a l a t e

O x a l a t e

Q u i n a t e

F r u c t o s e

G l u c o s e

S u c r o s e

C y a n i n

C y - 3 - O - S o p h

C y - 3 - O - S a m b

C y - G l u c - R u t

C y - 3 - O - G l u c

P e l - 3 - O - S o p h

C y - X y l - R u t

P e l - G l u c - R u t

C y - 3 - O - R u t

H y p e r o s i d e

Q u e r c G l u c

Citrate

0.88 0.65

-0.26

0.20

0.04

-0.07

-0.58

-0.25

-0.40

-0.58

-0.33

-0.55

-0.61

-0.54

-0.47 0.29

-0.19

Malate 0.88 0.91 -0.52 -0.22 -0.41 -0.08 -0.75 -0.49 -0.70 -0.87 -0.70 -0.81 -0.88 -0.85 -0.81 0.07 -0.33

Oxalate 0.65 0.91 -0.80 -0.34 -0.56 0.19 -0.73 -0.57 -0.74 -0.86 -0.80 -0.79 -0.85 -0.87 -0.86 -0.31 -0.62

Quinate -0.26 -0.52 -0.80 0.20 0.42 -0.61 0.54 0.54 0.54 0.49 0.63 0.48 0.46 0.57 0.61 0.81 0.90

Frucose 0.20 -0.22 -0.34 0.20 0.97 0.50 0.52 0.67 0.75 0.67 0.79 0.69 0.65 0.68 0.72 0.17 -0.17

Glucose

0.04

-0.41

-0.56

0.42

0.97 0.35

0.65

0.75 0.85

0.79

0.91 0.80

0.77

0.81

0.85

0.28

0.04

Sucrose -0.07 -0.08 0.19 -0.61 0.50 0.35 0.27 0.26 0.33 0.33 0.22 0.38 0.35 0.26 0.23 -0.68 -0.88

Cyanin -0.58 -0.75 -0.73 0.54 0.52 0.65 0.27 0.93 0.95 0.86 0.88 0.94 0.86 0.89 0.89 0.22 0.17

Cy-3-O-Soph -0.25 -0.49 -0.57 0.54 0.67 0.75 0.26 0.93 0.94 0.73 0.88 0.85 0.72 0.80 0.83 0.42 0.14

Cy-3-O-Samb -0.40 -0.70 -0.74 0.54 0.75 0.85 0.33 0.95 0.94 0.92 0.98 0.98 0.91 0.95 0.96 0.26 0.14

Cy -Gluc- Rut -0.58 -0.87 -0.86 0.49 0.67 0.79 0.33 0.86 0.73 0.92 0.93 0.98 1.00 0.99 0.98 0.03 0.15

Cy-3-O-Gluc -0.33 -0.70 -0.80 0.63 0.79 0.91 0.22 0.88 0.88 0.98 0.93 0.95 0.92 0.96 0.98 0.33 0.25

Pel-3-O- Soph -0.55 -0.81 -0.79 0.48 0.69 0.80 0.38 0.94 0.85 0.98 0.98 0.95 0.98 0.98 0.98 0.09 0.10

Cy-Xyl-Rut -0.61 -0.88 -0.85 0.46 0.65 0.77 0.35 0.86 0.72 0.91 1.00 0.92 0.98 0.99 0.97 -0.01 0.13

Pel-Gluc-Rut -0.54 -0.85 -0.87 0.57 0.68 0.81 0.26 0.89 0.80 0.95 0.99 0.96 0.98 0.99 1.00 0.15 0.22

Cy-3-O-Rut -0.47 -0.81 -0.86 0.61 0.72 0.85 0.23 0.89 0.83 0.96 0.98 0.98 0.98 0.97 1.00 0.22 0.25

Hyperoside 0.29 0.07 -0.31 0.81 0.17 0.28 -0.68 0.22 0.42 0.26 0.03 0.33 0.09 -0.01 0.15 0.22 0.76

Querc Gluc -0.19 -0.33 -0.62 0.90 -0.17 0.04 -0.88 0.17 0.14 0.14 0.15 0.25 0.10 0.13 0.22 0.25 0.76

O A

S u g a r s

LFsninaycohtnAsraguSAO

LFsninaycohtnAsraguSAO

O A

S u g a

r s

A n t h o c y a n i n s

F l


F

l

Fig.2. Correlationmatricesof phytochemical parameters ofraspberryfruit storedfor(A) 1 d and(B) 8 d at 2–3◦

C inthe dark.The correlationstatisticranges from−1 through0 to 1, indicating a ‘perfect negative correlation’, ‘no correlation’, and ‘perfect positive correlation’. Correlations of statistical significance (P

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Fig. 3. Venn-Diagram representing the number of unique metabolite–metabolitecorrelationsfor berries storedfor 1 or8 d andthe respective numberof correlationsthat were shared between the two storage treatments. Cut off point r −0.75.

berries stored for 8 d (r 0.05). In contrast, fructose showedstronger positive correlations (r >0.5) with all anthocyanins inberries stored for 8 d (Fig. 2B). The relationship between glu-cose and all anthocyanins remained similar for both 1 d and 8d stored berries. Only one significant correlation was observedbetween glucose and cyanidin-3-O-glucoside in samples storedfor 8 days. However, only very weak positive and non-significant

correlations between sucrose and anthocyanins were observed insamples stored for 8 d. Furthermore, sucrose and fructose onlyshowed weak positive and non-significant correlations with theflavonols hyperoside and quercetin-3-O-glucoside, while glucosewas strongly positively correlated with both flavonols after 1 d of storage (r >0.75, Fig. 2A). A different result was found for samplesstored for 8 d: While no correlation was found for fructose and glu-cose with either flavonol, a strong negative correlation was presentfor sucrose and hyperoside (r =−0.68, P > 0.05) and quercetin-3-O-glucoside (r =−0.88, P 0.75, P < 0.05) in samples storedfor 8 d in comparison to samples stored for only 1 d (Fig. 2).

All regression analysis results of individually quantified phy-tochemicals are summarised in Table 5. First, the relationshipbetween chemical compound concentration and maturity stagewas tested across the two storage times. Linear regression anal-ysis revealed that the slopes of all organic acids and anthocyaninswere significantly different from zero (P < 0.05), thus revealing alinear relationship between the concentrations of the respectiveorganic acids or anthocyanins and the maturity stage at harvest.Ontheotherhand,nosignificantlinearrelationshipswereobservedbetweenmaturitystageatharvestandtheconcentrationsofsugars,

or flavonols (Table 5).

In order to investigate if storage of 1 and 8 days influencedthe content of individual compounds, we set the regression slopesof each to be the same and tested if the intercept was sig-nificantly different. As can be seen from Table 5, significantdifferences in intercepts for the two storage treatments wereobserved for malate (P

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F r e s h n e s s

S w e e t n e s s

A c i d i t y

B i t t e r n e s s

T a s t e f u l n e s s

V i s u a l A t t r a c t i v e n e s s

S o l u b l e S o l i d s

T i t r a t a b l e A c i d s

S u g a r : A c i d R a t i o

Citrate 0.64 -0.22 0.52 -0.15 0.34 0.63 -0.47 0.81 -0.82Malate 0.25 -0.33 0.31 -0.10 0.14 0.38 -0.50 0.83 -0.80

Oxalate 0.62 -0.64 0.75 -0.83 -0.49 0.26 -0.08 0.14 -0.15

Quinate -0.13 0.25 -0.13 0.53 0.15 -0.08 0.45 -0.02 0.10

Frucose 0.13 -0.13 0.16 -0.62 -0.28 -0.15 -0.30 -0.22 0.17

Glucose -0.10 0.18 -0.13 -0.39 -0.14 -0.32 -0.25 -0.44 0.38

Sucrose -0.69 0.72 -0.88 0.52 0.36 -0.53 0.25 -0.60 0.59

Cyanin -0.36 0.53 -0.36 0.13 -0.08 -0.62 0.33 -0.80 0.78

Cy-3-O-Soph -0.05 0.22 -0.04 -0.18 -0.27 -0.38 0.27 -0.70 0.68

Cy-3-O-Samb -0.32 0.43 -0.32 -0.01 -0.17 -0.57 0.30 -0.83 0.80

Cy -Gluc- Rut -0.67 0.60 -0.69 0.21 -0.06 -0.76 0.34 -0.92 0.90

Cy-3-O-Gluc -0.59 0.55 -0.58 0.10 -0.08 -0.68 0.15 -0.78 0.75

Pel-3-O- Soph -0.47 0.21 -0.32 -0.20 -0.45 -0.76 0.28 -0.85 0.86

Cy-Xyl-Rut -0.70 0.62 -0.72 0.25 -0.03 -0.78 0.35 -0.91 0.89

Pel-Gluc-Rut

-0.76

0.43

-0.65

0.06

-0.25

-0.83

0.25

-0.86

0.86Cy-3-O-Rut -0.76 0.52 -0.70 0.15 -0.11 -0.76 0.17 -0.78 0.77

Hyperoside -0.40 0.28 -0.35 -0.04 -0.09 -0.24 -0.30 -0.35 0.27

Querc Gluc -0.32 0.21 -0.27 0.02 -0.09 -0.15 -0.20 -0.33 0.26

serocSlacimehCserocSyrosneS

O A

S u g a r s


F L

619

620

Fig. 4. Pearson’s correlation analysis of phytochemical components versussensory and chemical scores characteristics as determinedin raspberry fruits. Correlation is basedon both 1 d and8 d storage data compiled together.Significant (P

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the anthocyanins there was a similar but weaker correlation withsensory sweetness (Fig. 4).

The correlation analysis results shownin Fig.4 demonstratethata wide range of phytochemical components was significantly anddiversely correlated with one or more of the sensory traits. Promi-nent in this respect were sucrose andthe organic acids,which weresignificantly and oppositely correlated with several sensory traits,thus demonstrating a potential relationship of these componentsfor fruit taste in general. Also, the correlation results between awide range of sensory traits and oxalate which was present in thefruit in minor concentrations only was particularly surprising (Fig.4; Table 4). However, it should be kept in mind that oxalic acid is astrong acid with a particularly sharp taste. The results in Fig. 4 alsodemonstrate a strikingly similar relationship between the concen-trations of the various chemical components and the sensation of freshness, acidity and the visual attractiveness of the fruits, whichalso variedin parallel with the measured TA,but inversely with themeasured sugar:acid ratios.

We conclude that changes in raspberry fruit maturity, whichis closely related to changes in a number of phytochemical com-ponents, can be precisely assessed by visual observation of fruitcolour andby physical measurements of fruit/receptacleadherenceor fruit compression resistance. Furthermore, the storage capacityof the fruits are also determined by colour and maturity at the timeof harvest and thus colour assessment when related to a standard-ised colour system such as the Natural Colour System (NCS), willsecure picking of berries with optimum quality.

Acknowledgements

Thanks to Iren Lunde Knutsen and Sigrid Flatland for technicalassistance and to the staff at BAMAs head office that performedthe sensory evaluation. We gratefully acknowledge financial sup-port of this work by funding from the Regional Research Funds inNorway (RFF-Vestlandet) through Project ID: 203935, and from theResearch Council of Norway through project No. 234312/E50. Fur-thermore, the James Hutton Institute acknowledges funding by IVB

as part of the NSR program: ClimaFruit Project No. 35-05-09.

Appendix A. Supplementary data

Supplementary data associated with this article can be found,in the online version, at http://dx.doi.org/10.1016/j.scienta.2015.08.045.

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