Sea Buckthorn (HippophaÃ« rhamnoides ssp. rhamnoides) Berries in

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Sea buckthorn (Hippophaë rhamnoides ssp. rhamnoides) berries in Nordic environment: Compositional response to latitude and weather conditions Jie Zheng, Heikki P. T. Kallio, and Baoru Yang J. Agric. Food Chem., Just Accepted Manuscript • Publication Date (Web): 24 May 2016 Downloaded from http://pubs.acs.org on May 24, 2016

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Journal of Agricultural and Food Chemistry

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Sea buckthorn (Hippophaë rhamnoides ssp. rhamnoides)

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berries in Nordic environment: Compositional response to

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latitude and weather conditions

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Jie Zheng a,b, Heikki Kallio a,b and Baoru Yang a,*

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a

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20014 Turku, Finland;

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b

Food Chemistry and Food Development, Department of Biochemistry, University of Turku, FI-

Department of Food Science and Engineering, Jinan University, 510632 Guangzhou, China

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Running title: Genotype, latitude and weather affect sea buckthorn composition

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*

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Turku, Finland. Tel.: +358 2 333 6844; Fax: +358 2 231 7666.

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Corresponding author. Address: Department of Biochemistry, University of Turku, FI-20014

E-mail address: [email protected] (B. Yang)

ACS Paragon Plus Environment

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ABSTRACT

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Flavonol glycosides (FGs) in sea buckthorn (Hippophaë rhamnoides ssp. rhamnoides) berries of

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varieties ‘Tytti’ and ‘Terhi’, cultivated in northern Finland (68°02′ N) for six years and southern

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Finland (60°23′ N) for seven years, were investigated and compared by HPLC-DAD-ESI-MS/MS.

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The average total content of 23 identified glycosides of isorhamnetin and quercetin was 103 ± 23

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and 110 ± 21 mg/100 g fresh berries in ‘Terhi’ and ‘Tytti’, respectively. The total contents of

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FGs, flavonol di-glycosides and tri-glycosides in both varieties were higher in north than in south,

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whereas total flavonol mono-glycosides content behaved vice versa (p < 0.05). Among the 89

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weather variables studied, the sum of the daily mean temperatures that are 5 °C or higher from

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the start of growth season until the day of harvest was the most important variable which

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associated negatively with the accumulation of FGs in berries. Such influence was much stronger

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in berries from north than from south.

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Keywords: flavonol, Hippophaë rhamnoides, latitude, sea buckthorn, variety, weather conditions


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Introduction Sea buckthorn (Hippophaë rhamnoides L.) plant is used for soil, water and wildlife

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conservation and anti-desertification purposes because of its strong roots with nitrogen fixing

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ability as well as its resistance to extreme conditions such as drought, cold, and salinity.1

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Because of their high nutritive value, the berries are widely consumed as food and food

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supplements in Europe and Asia.2 Sea buckthorn berries are rich in flavonol glycosides with

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characteristic composition and properties. Isorhamnetin is the typical and most abundant

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aglycone, and low quantities of quercetin glycosides are also found in the berries.3-5

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Isorhamnetin glycosides (IGs) are less common in fruits and berries than the glycosides of

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kaempferol, quercetin and myricetin.6 Isorhamnetin derivatives have been found in lesser

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amounts in e.g. apples,7 onions8 and raspberries.9 However, Jiménez-Aspee et al.10 reported

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isorhamnetin-3-O-rutinoside to be the major phenolic compound in copao fruits (Eulychnia

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acida Phil., Cactaceae).

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Use of isorhamnetin is patented in China and the compound is used as an active ingredient for

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preparing medicine or health foods for treating enteritis and ulcerative colitis.11 It was reported

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that the anticoagulant and profibrinolytic effects of isorhamnetin-3-O-galactoside were greater

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than those of quercetin-3-O-galactoside.12 This indicates positive enhancement of the

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anticoagulant function by the methoxy group of isorhamnetin-3-O-galactoside. Yang et al.13,14

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reported the potential of isorhamnetin in inhibition of the acute inflammatory response and

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protection of hepatocytes against oxidative stress. Antunes-Ricardo et al.15 reported isorhamnetin

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glycosides isolated from Opuntia ficus-indica pads to show cytotoxic effect against colon cancer

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cells of HT-29 and Caco2 in vitro. Moreover, isorhamnetin di-glycosides were reported to be

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more cytotoxic than pure isorhamnetin aglycone or tri-glycosides when they were tested with


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HT-29 cells. The research demonstrated that glycosylation has a significant effect on the

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antiproliferative effect of isorhamnetin glycosides. The significant health-maintaining properties

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of isorhamnetin derivatives and their potential usage as ingredients in many health foods and

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remedies brought special interests in the research of isorhamnetin glycosides in sea buckthorn.

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Previous studies showed that genetic background, cultivation methods, harvesting time,

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growth sites, and weather conditions all have impacts on the accumulation of primary and

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secondary metabolites in fruits and berries.16-22 Accumulation of the secondary metabolites is

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triggered under various unfavorable environmental stresses. For instance, exposure of plant to

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elevated UV-B light has been reported to enhance the accumulation of flavones and flavonols.23

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Nordic latitudes at Arctic Circle (66°33′46″N) and beyond provide special stresses for plants to

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grow and might cause abundant accumulation of secondary metabolites for defense against the

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environmental stresses. This research focuses on the impact of growth latitude and weather

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conditions on the accumulation of flavonol glycosides in sea buckthorn berries of different

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varieties suitable for cultivation in the far north.

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During the past years, our research group has cultivated nine different varieties of sea

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buckthorn bushes in southern and northern Finland, covering a latitudinal distance of 850 km:

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‘Avgustinka’, ‘Botanicheskaya’, ‘Trofimovskaya’, ‘Pertsik’, ‘Prevoshodnaya’,

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‘Prozcharachnaya’, ‘Raisa’, ‘Terhi’ and ‘Tytti’. Only three varieties, ‘Terhi’, ‘Tytti’ and ‘Raisa’

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survived and produced fruits after acclimation to the extreme conditions in northern Finland. The

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cultivar ‘Raisa’, with clearly lower crop yield, is not as promising as ‘Tytti’ and ‘Terhi’. ‘Terhi’

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and ‘Tytti’ stand out as the most suitable varieties for cultivation at high latitudes with extreme

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weather conditions.


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In the present study, cultivated sea buckthorn berries of varieties ‘Terhi’ and ‘Tytti’ were

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harvested in Sammalmäki in southern Finland and in Kittilä in northern Finland over seven years.

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The effects of latitude and weather conditions on the compositional profile of flavonol

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glycosides were studied thoroughly. Moreover, the compositional differences between samples

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of different varieties, and the correlation between the metabolites in berries were studied.

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Materials and methods

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Samples

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Sea buckthorn (Hippophaë rhamnoides ssp. rhamnoides) of varieties ‘Terhi’ and ‘Tytti’ were

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planted at two different growth locations in Finland to investigate the effects of growth latitude

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and weather conditions on the composition of flavonol glycosides (FG) of the berries. Seedlings

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were planted in Sammalmäki, Turku, Finland (longitude 22°09′ E, latitude 60°23′ N, altitude 1 m)

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in 2004 and in Kittilä, Finland (24°37′ E, 68°02′ N, 210 m) in 2002–2003. Berries were picked

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optimally ripe and loosely frozen at –20 °C immediately after picking. In order to overcome the

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plant-to-plant variation, the berries were harvested separately from several bushes in both growth

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places. The berries picked from one bush were treated as one individual sample lot. Four sample

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lots were harvested annually in Sammalmäki during 2007–2013 and five sample lots in Kittilä

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during 2008–2013. However, the berries collected in Kittilä in 2008 were considered half-ripe

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according to their appearance, and were excluded from the statistical analysis.

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Chemicals

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Quercetin-3-O-rutinoside, quercetin-3-O-glucoside, isorhamnetin-3-O-glucoside and

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isorhamnetin-3-O-rutinoside were purchased from Extrasynthese (Genay, France). Reference

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compounds of quercetin-3-O-sophoroside-7-O-rhamnoside, isorhamnetin-3-O-glucoside-7-O-

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rhamnoside and isorhamnetin-3-O-sophoroside-7-O-rhamnoside were provided by our


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collaborators. 24 Methanol, tetrahydrofuran, trifluoroacetic acid and acetonitrile were of HPLC

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grade or MS grade, or the highest grades available.

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HPLC-DAD analysis of flavonol glycosides

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The flavonol glycosides were extracted from sea buckthorn berries and analyzed with an

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HPLC-DAD system in quadruplicate in the same way as described by Ma et al.24 Flavonol

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glycosides were detected at 360 nm, and quantified by the external standards curves of quercetin-

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3-O-glucoside, quercetin-3-O-rutinoside, isorhamnetin-3-O-glucoside and isorhamnetin-3-O-

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rutinoside. Quercetin-3-O-glucoside and isorhamnetin-3-O-glucoside were used as the external

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standard for the glucosides and for all the mono-glycosides of each aglycone, respectively.

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Quercetin-3-O-rutinoside and isorhamnetin-3-O-rutinoside were used as the external standard for

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the rutinosides, and for all the di-glycosides and tri-glycosides of each aglycone, respectively.

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Identification of flavonol glycosides by HPLC-DAD-ESI-MS/MS

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The identification of flavonol glycosides was based on UV spectra, retention times, reference

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compounds, mass spectra, and literature data.3,4,25 The samples were prepared as described above

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and filtered through a 0.2 µm filter. The HPLC-MS system and conditions were the same as

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described by Ma et al with modifications.24 A Phenomenex Aeris Peptide XB-C18 column (250

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× 4.60 mm i.d., particle size 5 µm) (Torrance, CA) was used for the analysis. For identification

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of different flavonol glycosides, two eluting gradient programs were applied. One of them was:

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0–2 min, 13% B; 2–14 min, 13–25% B; 14–19 min, 25% B; 19–24 min, 25–60% B; 24–28 min,

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60% B; 28–30 min, 60–90% B; 30–35 min, 90% B; 35–40 min, 90–13% B; 40–50 min, 13% B.

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The other one was: 0–5 min, 13% B; 5–20 min, 13–25% B; 20–25 min, 25% B; 25–30 min, 25–

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60% B; 30–34 min, 60% B; 34–36 min, 60–90% B; 36–41 min, 90% B; 41–50 min, 90–13% B;

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50–60 min, 13% B. The flow rate of the mobile phase was 0.5 mL/min.


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Information of weather conditions

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The weather information was obtained from the Finnish Meteorological Institute (Helsinki,

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Finland). The data were recorded at the weather station closest to the collecting points in

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Artukainen, Turku (22°10'E, 60°27'N, 8 m) and Pokka, Kittilä (25°47'E, 68°10'N, 275). The data

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were also recorded in Kaarina Yltöinen (22°33'E, 60°23'N, 6 m) and Sodankylä (26°37'E,

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67°21'N, 179 m) as a back-up for the data collection in Artukainen, Turku and Pokka, Kittilä,

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respectively. The abbreviations of all the weather variables are listed in Table 1. SUMTgs is the

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sum of the daily mean temperatures that are 5 °C or higher during the growth season. It is

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calculated using the following formula: SUMTgs= ∑ Ti ( Ti ≥ 5 ), where Ti is the daily mean

b i =a

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temperature on day i, and a is the start of growth season and b is the last day of growth season.

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SUMTgh is the sum of the daily mean temperatures that are 5 °C or higher from the start of

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growth season until the day of harvest. It is calculated using the following formula:

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SUMTgh= ∑ Ti ( Ti ≥ 5 ), where Ti is the daily mean temperature on day i, and c is the start of

d i =c

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growth season and d is the day of harvest. SUMTm is the sum of the daily mean temperatures

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that are 5 °C or higher in the last month before harvest. It is calculated using the following

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formula: SUMTm= ∑ Ti ( Ti ≥ 5 ), where Ti is the daily mean temperature on day i, and e is the

f

i =e

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30 days before harvest and f is the day of harvest.

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Statistical analysis

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Statistical analyses were performed with SPSS 22.0.0.1 (SPSS, Inc., Chicago, IL) and

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Unscrambler 10.3 (Camo Process AS, Oslo, Norway). The samples of ‘Terhi’ and ‘Tytti’

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collected from Kittilä in 2008 were excluded because of the half-ripe berries. Independent-


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samples t-test was applied to study the compositional difference between sea buckthorn varieties

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‘Terhi’ and ‘Tytti’, and between berries grown at different locations. PLS-DA (Partial Least

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Squares - Discrimination Analysis) was applied to investigate the difference in the composition

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of berries between different growth locations.

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The impact of weather variation on the berry composition was studied by principal component

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analysis (PCA) and Pearson’s correlation coefficients analysis. To simplify the PCA plots,

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weather variables including the maximum and minimum temperatures in different months were

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eliminated from the principal component analysis. Correlations between these parameters and

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compositional parameters of the berries were available via Pearson’s correlation coefficients

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(Table S1–S6). The correlations between different metabolites were investigated with Pearson’s

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correlation coefficients analysis as well in this study.

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Results and discussion

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Identification of flavonol glycosides

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Figure S1 shows the HPLC-DAD chromatograms of flavonol glycosides in sea buckthorn

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berries of varieties ‘Tytti’ and ‘Terhi’. In total, 23 compounds were either unambiguously or

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tentatively identified (Table S7). Quercetin-3-O-sophoroside-7-O-rhamnoside (Q-3-S-7-Rh),

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isorhamnetin-3-O-sophoroside-7-O-rhamnoside (I-3-S-7-Rh), isorhamnetin-3-O-glucoside-7-O-

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rhamnoside (I-3-G-7-Rh), quercetin-3-O-rutinoside (Q-3-R), quercetin-3-O-glucoside (Q-3-G),

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isorhamnetin-3-O-rutinoside (I-3-R) and isorhamnetin-3-O-glucoside (I-3-G) were

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unambiguously identified through the retention times, UV spectra and mass spectra of the

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reference compounds, and by co-elution with standards. These seven FGs covered 79 % and

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70 % of all the FGs in the berries of ‘Terhi’ and ‘Tytti’, respectively.


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In the current study, hexose (m/z 162) and deoxyhexose (m/z 146) were the only sugar

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moieties detected in flavonol glycosides. In the study conducted by Ma et al.24, also pentosides of

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flavonols were found in sea buckthorn of subspecies mongolica and sinensis. Rösch et al.

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reported glucose and rhamnose to be the exclusive sugars detected via HPTLC analysis after acid

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hydrolysis in sea buckthorn (Hippophaë rhamnoides ssp. rhamnoides) pomace.25 Therefore,

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compounds 2, 4, 7, 12, 14, 16 and 17 were tentatively identified as glycosides of quercetin or

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isorhamnetin attached with different sugar moieties of glucose and rhamnose. Compound 7, 12,

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14, 16 and 17 were defined without detailed information of the positions of the sugar attachments

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(Table 2). These compounds formed 15 % and 22 % in the total FGs in ‘Terhi’ and ‘Tytti’,

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respectively. Thus, > 90 % of all the FGs was identified at the level of aglycones and sugar

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moieties. Although appearance of glycosides of kaempferol and myricetin in sea buckthorn

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berries has been reported in other studies,4,5,25-27 they were not detected in the current study.

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Due to the low concentration in the berry samples, compounds 5 and 6 could be only

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tentatively identified as quercetin glycosides with the clear aglycon pseudomolecular ion at m/z

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303 in the mass spectra. However, quantitative analysis was not possible because of the co-

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elution of unknown compounds in some of the samples. Because of the co-elution, peaks of

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isorhamnetin-glucoside-rhamnoside 2 (compound 12) and quercetin-3-O-glucoside (compound

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13) were integrated together and quantified as quercetin-3-O-glucoside. Compounds 9, 10, and

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19–23 were identified as isorhamnetin glycosides with the aglycon pseudomolecular ion at m/z

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317. Considered as minor compounds, they were quantified as a group of unknown isorhamnetin

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glycosides.

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Flavonol glycoside profiles and comparison of the varieties


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Isorhamnetin glycosides were the main FGs in berries of varieties ‘Terhi’ (88.9% of total

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FGs) and ‘Tytti’ (86.6%) (Table 2). This is in accordance with the results reported previously in

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other studies on the same subspecies ssp. rhamnoides as well as on the subspecies sinensis,

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mongolica, carpatica, yunnanensis and wolongensis.3-5 However, there are differences in

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compositional profiles with varying proportions of individual flavonols among different

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subspecies and varieties.3-5 Chen et al. reported I-3-G-7-Rh to be the most abundant FG in H.

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rhamnoides ssp. sinensis and ssp. yunnanensis, while I-3-R was the most abundant one in ssp.

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wolongensis. In contrast to the findings of Chen et al., Yang et al.3 reported I-3-R to be the most

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abundant FG in berries of ssp. sinensis from Wenshui and Xixian, China. In the current study, I-

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3-R (29.4% of total FGs) was the most abundant FG in ‘Terhi’, followed by I-3-G-7-Rh (24.9%)

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and I-3-G (9.0%). Again, in the variety ‘Tytti’, I-3-G-7-Rh (23.9%) was detected to be the most

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abundant FG, with I-3-R (16.6%) and I-3-G (11.3%) being the second and third most abundant

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ones. Isorhamnetin di-glycosides (I-digly) accounted for 54.2–62.6% and dominated in total

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flavonols in all the sea buckthorn samples studied.

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In half-ripe berries from Kittilä in 2008 the total content of FGs was double when compared

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with ripe berries (Table 2). I-3-G-7-Rh and I-3-R represented 31.2–31.7% and 17.2–29.2% of

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total FGs in the half-ripe berries, giving an indication of accumulation and metabolism of

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flavonols during ripening. A common knowledge is that flavonols typically peak in unripe fruits

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and decrease during ripening.28-30

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The total content of FGs was lower in ‘Terhi’ than in ‘Tytti’, 103 mg/100 g vs.110 mg/100 g

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fresh berries, respectively. The small difference between the varieties was, however, statistically

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significant (p < 0.05). It has been reported that FGs of different aglycones or of different

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attachments of sugar moieties exhibited divergent/dissimilar antioxidant capacity, bioavailability


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and heath-beneficial effects.12,15,31,32 Quercetin and quercetin glycosides show higher antioxidant

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activity than glycosides of isorhamnetin and kaempferol,32 while isorhamnetin-3-O-galactoside is

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reported to exhibit greater anticoagulant and profibrinolytic effects than those of quercetin-3-O-

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galactoside.12 Therefore, a more specific comparison of individual FGs is necessary.

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Isorhamnetin glycosides (IGs) comprised the majority (> 85 %) of FGs in both varieties studied.

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The total content of IGs was identical (p > 0.05) between the two varieties, being 91.8 mg/100 g

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in ‘Terhi’ and 95.4 mg/100 g in ‘Tytti’, (Table 2). The highest varietal difference (by 66%, p

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0.05, Table 2). The total content of isorhamnetin di-glycosides was 8 % higher (p < 0.05) in

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‘Terhi’ than in ‘Tytti’, whereas mono-glycosides and tri-glycosides were 27% and 21% lower in

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‘Terhi’ than in ‘Tytti’, respectively. In the study conducted by Antunes-Ricardo et al.15,

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isorhamnetin di-glycosides showed more cytotoxic effect against colon cancer cells of HT-29

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than pure isorhamnetin aglycone or tri-glycosides in vitro.

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Quercetin glycosides (QGs) comprised 10-15 % of total FGs. All the four individual QGs

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identified in this study showed significantly higher contents in ‘Tytti’ than in ‘Terhi’ (Table 2).

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Thus, low but constantly significant differences in both IGs and QGs between the two varieties

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were observed over years. When comparing the berries at each orchard (in Sammalmäki and in

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Kittilä, respectively) separately, the same result of varietal comparison was observed.

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Latitudinal comparison

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PLS-DA was applied to create predictive models for sea buckthorn of both varieties to

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differentiate samples collected from the two growth locations based on berry composition. As a


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result, clear separation (R2 = 80.2% and 87.2%, and Q2 = 79.5% and 87.0% for ‘Terhi’ and

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‘Tytti’, respectively) of samples from different growth sites were observed in the score plots of

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Figure 1.

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‘Terhi’ and ‘Tytti’ showed similar compositional response to the growth latitude. Sea

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buckthorn samples cultivated in Kittilä in Finnish Lapland, north of the Polar Circle was clearly

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separated from those grown in Sammalmäki in southern Finland. According to the loading plots

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of ‘Terhi’ and ‘Tytti’, Kittilä and Sammalmäki were highly explained and separated by both

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Factor 1 and Factor 2. Most of the flavonol glycosides, such as I-3-S-7-Rh, Q-3-S-7-Rh, I-3-G-7-

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Rh, Q-3-G-7-Rh, I-3-R, Q-3-R, I-G-Rh 1 and I-G-Rh 3, were all explained by Factor 1 and

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showed significantly higher values (by 25.0–117.5% and 26.5–97.2% in ‘Terhi’ and ‘Tytti’,

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respectively) in berries from northern Finland than those from southern Finland. As a result, the

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total contents of di-glycosides of isorhamnetin (I-digly) and quercetin (Q-digly) , tri-glycosides

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of isorhamnetin (I-trigly), quercetin glycosides, isorhamnetin glycosides, and flavonol glycosides

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were significantly higher (by 30.5–52.4% in ‘Terhi’ and 18.8–60.7% in ‘Tytti’, respectively) in

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berries from Kittilä than from Sammalmäki. In contrast, the total content of isorhamnetin

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glucosides (compounds 13 and 17, the only isorhamnetin monoglycosides), was highly explained

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by Factor 2 in both varieties and showed higher values (by 44.6% and 104.2% in ‘Terhi’ and

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‘Tytti’, respectively) in berries grown in Sammalmäki than those in Kittilä (Figure 1). More

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specifically, I-3-G was more abundant in berries from south, by 55.4% in ‘Terhi’ and 119.6% in

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‘Tytti’, than in north. The other two minor mono-glycosides, I-G and Q-3-G exhibited the same

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trend in ‘Tytti’, while no significant difference between growth locations in ‘Terhi’ was observed.

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Also the isorhamnetin-glucoside-rhamnoside 4, located close to Sammalmäki in the loading plots


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of both Terhi and Tytti, were 2-3 times more abundant in berries from the South than those from

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the North.

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The weather at the higher latitude was generally characterized with lower temperature and

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light intensity when compared to the weather at the lower latitude. Schulz et al.33 studied the

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impact of cold acclimation on the variation of flavonol and anthocyanin metabolism in plant

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Arabidopsis thaliana. They found that the content of flavonoid increased after cold acclimation

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(4 °C) for two weeks, which suggested the important role of post-transcriptional mechanism in

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the regulation of flavonoid metabolism under cold conditions. However, the environmental

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conditions in the current investigation were complex and different from their study. More

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detailed investigations into the effects of weather conditions on the composition of the berries is

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requested to dig out the actual factors affecting metabolism and final composition of sea

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buckthorn berries.

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Effects of environmental factors

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Principal component analysis (PCA) was applied to provide an overview on the compositional

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response of sea buckthorn berries from different growth locations and harvesting years to the

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weather conditions, separately for ‘Terhi’ and ‘Tytti’ (Figure 2). In the PCA plots, the first two

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principal components (PCs) explained 94% of the data variance of berry composition and

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weather conditions of both varieties, and they behaved quite similarly in view of the response to

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the weather conditions. PC1 explained 91 % of the variance for both ‘Terhi’ and ‘Tytti’, and

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separated the compositional parameters especially based on their response to temperature and

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radiation. Pearson’s correlation coefficients (r) related to the parameters shown in Figure 2 are

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collected in Supplementary materials as Table S1–S6.


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Several major FGs, e.g. I-3-G-7-Rh, I-3-S-7-Rh, Q-3-S-7-Rh, Q-3-G-7-Rh, I-3-R, I-G-Rh 1,

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as well as I-digly, Q-digly and I-trigly showed negative correlations with the major temperature

284

and radiation parameters, in both ‘Terhi’ and ‘Tytti’ (Table S1 and S4). They were all well

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explained by PC1 and located oppositely far away from the temperature and radiation parameters

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(Figure 2). In contrast, I-3-G, I-G-Rh 4 and total isorhamnetin mono-glycosides (I-monogly) had

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positive correlations with the temperature and radiation parameters in both berry varieties. This

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indicates the central role of temperature or sunshine in degradation or transformation of FGs into

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more simple components. The content of the minor I-3-R-7-G seemed be hardly effected by the

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weather conditions. Among the radiation parameters, the monthly solar irradiance in March,

291

April, May, July and August, compared to the others, were weakly explained by PC1and showed

292

less influence on regulation of accumulation of FGs (Table S1 and S4).

293

In accordance with the response of the major FGs towards the variation of temperature and

294

radiation, total quercetin glycosides (Total QG), total isorhamnetin glycosides (Total IG) and

295

total flavonol glycosides (Total FG) were negatively correlated with temperature and radiation

296

variables. However, the explanation of Total IG and Total FG by PC1 in the loading plots was

297

much weaker in ‘Tytti’ than in ‘Terhi’ (Figure 2), which indicated weak correlations with

298

temperature and radiation variables in ‘Tytti’ (Table S4). The total content of unknown

299

isorhamnetin-glycosides (Unk I-gly) showed clear positive correlations with temperature and

300

radiation variables only in Tytti.

301

In contrast to the current study, the total content of flavonol glycosides in black currant

302

varieties ‘Mortti’ and ‘Ola’ displayed positive associations with the temperature and radiation

303

variables.16 In the other currant varieties of ‘Melalahti’,16 ‘Red Dutch’, ‘White Dutch’ and

304

‘Vertti’,19 the contents of flavonol glycosides were less dependent on the radiation and


14

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305

temperature variables. This indicates differences in the regulation of flavonoid metabolism

306

between different plant materials in response to the variation of weather conditions. However,

307

the correlation between weather conditions and other phenolic compounds, eg. hydroxycinnamic

308

acid conjugates and anthocyanins, in currant berries is worth to be noticed.

309

Compared to temperature and solar radiation, precipitation and humidity had less influence on

310

the composition of both varieties. All the precipitation variables were located close to the center

311

of the plots, indicating no effects on the contents of FGs. Humidity variables, again, showed

312

better correlation with the phenolics investigated. Although most of the humidity variables

313

exhibited no correlation with flavonol composition, some specific variables showed influence on

314

accumulation of certain components (Figure 2). The average humidity from the start of growth

315

season until the day of harvest (Hgh) and in the last month before harvest (Hm), percentage of

316

the days with a relative humidity of 90–100% from the start of growth season until the day of

317

harvest (DH90to100gh) and in the last month before harvest (DH90to100m) showed positive

318

influence on accumulation of I-3-S-7-Rh, Q-3-S-7-Rh, I-3-G-7-Rh, Q-3-G-7-Rh, and I-G-Rh 1

319

(Table S1 and S4). Thus, berries typically tend to accumulate more of these compounds under

320

extremely high humidity. Again, variables like average humidity in January (Hjan) and February

321

(Hfeb), percentage of the days with a relative humidity of 60–70% from the start of growth

322

season until the day of harvest (DH60to70gh) and percentage of the days with a relative

323

humidity of 70–80% in the last month before harvest (DH70to80m) showed negative correlations

324

with the flavonol glycosides mentioned above (Figure 2, Table S1 and S4). I-G-Rh 4, in contrast

325

to the flavonols mentioned above, exhibited positive correlations with Hjan, Hfeb, DH60to70gh

326

and DH70to80m, and negative correlation with Hgh, Hm, DH90to100gh and DH90to100m. It


15


327

indicates again, as in previous researches,16,34 that the regulation of metabolic and biosynthetic

328

pathways in berries may respond to weather variables differently at different growth periods.

329

Page 16 of 36

The correlation loadings shown in Figure 2 were complemented with more detailed analysis

330

of dependence of individual FGs on weather conditions (Figure 3). The sufficient number of

331

years of investigation (five in Kittilä and seven in Sammalmäki) made it possible to get a

332

generalized view of the climatic effects. It was indicated by Figure 3 that the composition of

333

berries depends widely on the growth site, year and weather conditions. Differences between

334

FGs from different places and different years could be as high as five-fold as e.g. in the case of I-

335

3-G-7-Rh. Concerning the big deviation between the climatic data from southern Finland and

336

northern Finland, the dependence of flavonol accumulation on the weather conditions were also

337

studied by Pearson’s correlation coefficient analysis in each growth site (Table S2, S3, S5 and

338

S6).

339

Figure 3A summarizes one of the common features of FGs in the berries studied, i.e. the clear

340

decreasing linear trend in contents of the major compounds I-3-G-7-Rh, I-3-S-7-Rh, I-3-R, and

341

Q-3-S-7-Rh in berries grown in Kittilä by an increase in the sum of the daily mean temperatures

342

that are 5 °C or higher from the start of growth season until the day of harvest (SUMTgh). Their

343

decreasing trends were in close accordance with the overall linear trends (Figure 3A). ‘Terhi’

344

and ‘Tytti’ behaved quite analogously which is a message of their genetic similarities. However,

345

some exceptions may exist. For example, a clear decreasing trend of Q-3-G-7-Rh towards an

346

increase in SUMTgh was only observed in berries of ‘Terhi’ but not the berries of ‘Tytti’ in

347

Kittilä. In berries from Sammalmäki, the same but weak decreasing trends of I-3-G-7-Rh, I-3-S-

348

7-Rh, Q-3-G-7-Rh and Q-3-S-7-Rh against the increasing values of SUMTgh were observed in

349

both ‘Terhi’ and ‘Tytti’. But the content of major flavonol I-3-R in berries from Sammalmäki


16

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350

was less affected by SUMTgh in both ‘Terhi’ and ‘Tytti’. These five FGs comprised 67 % (69

351

mg/100 g) of all the FGs in ‘Terhi’ and 55 % (60 mg/100 g) in ‘Tytti’.

352

It is widely known that the synthesis of flavones and flavonols in plants are generally triggered

353

by elevated UV-B light.23 However, temperature was another factor influencing the

354

accumulation of flavonols. Strawberry cultivars ‘Earliglow’ and ‘Kent’ showed an increase as

355

high as ten-fold in the contents of flavonols in response to the increase of temperature from

356

18/12°C to 30/22°C (day/night)35 which is in complete contrast to our findings. In the current sea

357

buckthorn study, flavonols were extracted from the whole berry, including both skin and pulp.

358

Pereira et al.36 suggested in the study carried out on grapes that the regulation of flavonol

359

synthesis by light and temperature was different in different parts of fruit, and even for different

360

compounds. The content myricetin-3-glucoside was higher in the pulp but lower in the skin of

361

shaded berries than sun-exposed ones. They proposed temperature to be the major factor

362

influencing the flavonol accumulation in grape pulp while light in grape skin. In contrast to

363

myricetin-3-glucoside, the contents of kaempferol-3-glucoside and quercetin-3-glucoside in the

364

pulp of sun-exposed berries were higher than those of shaded berries, which suggested that

365

different flavonols responded to environmental conditions differently.

366

Development of I-3-G-7-Rh, I-3-S-7-Rh, I-3-R, Q-3-G-7-Rh and Q-3-S-7-Rh on two other

367

temperature sum parameters, the sum of the daily mean temperatures that are 5 °C or higher

368

during the growth season (SUMTgs) and in the last month before harvest (SUMTm) are shown

369

in Figures 3B and 3C, respectively. The linear profiles in Sammalmäki and Kittilä deviate

370

typically more from the overall trends when compared with Figure 3A. Especially temperature

371

conditions in the last month before harvest had almost no effect on the five selected FGs in

372

Sammalmäki (Figures 3C).


17


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373

When the correlations between flavonol glycosides and some other weather variables were

374

investigated in Sammalmäki and Kittilä separately, the linearity was lost in many compounds, or

375

it showed even opposite direction compared with results when the two locations were combined.

376

An interesting example of effects of average temperature in February (Tfeb) is shown in Fig 3D.

377

The lower the temperature was in south Finland in February, the lower were the contents of I-3-

378

G-7-Rh, I-3-S-7-Rh, I-3-R, Q-3-G-7-Rh and Q-3-S-7-Rh in the berries in next autumn, in both

379

‘Terhi’ and ‘Tytti’. In Kittilä the correlations between the contents of these compounds and Tfeb

380

were less evident than in Sammalmäki. In addition to the temperature variables, specific

381

correlations between certain FGs and variables of radiation, humidity and precipitation were also

382

detected (Figure 3E). Again, big deviations were observed between berries grown at different

383

sites in the compositional response of FGs to the variation of these weather variables.

384

The harvesting dates were not selected by calendar but defined by sensory properties

385

evaluated by an experienced person. All the samples were collected over the years within a frame

386

of three weeks in autumn. It was observed that in Kittilä the slower the accumulation of the

387

temperature parameters were, i.e. the lower the values of SUMTgs, SUMgh and SUMTm in the

388

corresponding year, the later the berries ripened, and the later harvesting took place (Figure S2).

389

But in Sammalmäki, the collection of ripe berries was independent of such temperature

390

parameters. In addition, clear increasing trends were observed in contents of the selected five

391

FGs, i.e. I-3-G-7-Rh, I-3-S-7-Rh, I-3-R, Q-3-G-7-Rh and Q-3-S-7-Rh, as the harvesting date of

392

berries delayed in Kittilä but no clear trends were detected in Sammalmäki (Figure S3), with the

393

only exception of Q-3-G-7-Rh in ‘Tytti’.

394

The content and composition of phenolic compounds in berries is commonly influenced by the

395

stage of ripeness.28-30 As the composition of Kittilä berries is very sensitive and dependent on the


18

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396

date of harvest defined according to ripeness, it was suggested by the results that even though the

397

berries were tried to be harvested at the same ripeness stage according to the appearance and

398

taste, deviations in ripeness may still exists. Unlike the berries in Sammalmäki which were

399

typically harvested before the end of the growth season, the berries in Kittilä ripen typically very

400

late and are mostly collected after the end of the growth season. Berries grown in the far north in

401

Finland were always smaller than those in the southern Finland.

402

According to the results obtained, it is possible to conclude that the majority of flavonol

403

contents are largely influenced by accumulation of temperature parameters especially by

404

SUMTgh. The effect was strongest in berries in Kittilä, mostly due to the dependence of ripeness

405

and harvest day on temperature sum parameters. When the temperature sum variables reached a

406

high level comparable to southern Finland, the content of flavonols was no longer affected by the

407

harvest date and its correlation with temperature sum parameters was much weaker than berries

408

in average grown in north. The Total FG showed significant negative correlations with SUMTgs,

409

SUMTgh and SUMTm in berries from Kittilä (r = –0.61 to –0.65 and –0.65 to –0.68 for ‘Terhi’

410

and ‘Tytti’, respectively, p < 0.01) but no clear correlation was observed in berries from

411

Sammalmäki (Table S1－－S6). The results suggest that heat absorption is essential for

412

degradation of flavonols, and the accumulation/degradation reaches a plateau when sufficient

413

amount of heat is absorbed by the plant, i.e. the accumulation of temperature reached a specific

414

level. However, more detailed investigations including recording of various wave lengths beyond

415

the visible light during the growth period should be conducted for more accurate conclusions.

416

Correlation between metabolites

417 418

In both sea buckthorn varieties studied, positive correlations between 3-O-sophoroside-7-Orhamnoside of quercetin and isorhamnetin (r = 0.94–0.97, p < 0.01), between glucosides of


19


Page 20 of 36

419

quercetin and isorhamnetin (r = 0.59–0.95, p < 0.01), and between rutinosides of quercetin and

420

isorhamnetin (r = 0.63–0.77, p < 0.01) were detected. In addition, Q-3-G-7-Rh, I-3-O-G-7-Rh

421

and I-G-Rh 1 showed positive correlations with each other (r = 0.85–0.98, p < 0.01), as well as

422

with I-3-S-7-Rh and Q-3-S-7-Rh (r = 0.81–0.93, p < 0.01). In contrast, I-G-Rh 4 displayed

423

negative correlations with I-3-S-7-Rh and Q-3-S-7-Rh (r = –0.74 to –0.83, p < 0.01). These

424

results indicated close relations between these metabolites in the biosynthetic pathways of

425

flavonoids in plants and provided useful information for the further physiological and genomic

426

investigations in sea buckthorn.

427

Funding

428

The work was financed by the Finnish Graduate School on Applied Bioscience:

429

Bioengineering, Food and Nutrition, Environment (ABS); the Turku University Foundation,

430

Finland; the Finnish Food Research Foundation, Finland, KAUTE foundation/Eeva-Liisa

431

Hirvisalo Fund, Finland; the Scholarship Fund of Pharmacist Wäinö Edvard Miettinen, Finland;

432

Alfred Kordelinin Säätiö, Finland; Jenny ja Antti Wihurin rahasto, Finland.

433

Acknowledgements

434

We are grateful to Hannu Lappalainen, Bärtil Lappalainen and Seppo Lappalainen for

435

providing the sea buckthorn berries over years for the study. We also acknowledge the technical

436

assistance by Pengzhan Liu and Oskar Laaksonen in the analysis.

437

Supporting Information

438

Pearson’s correlation coefficients between weather conditions and compositional parameters of

439

sea buckthorn berries (Table S1–S6), identification of flavonol glycosides (Table S7), HPLC

440

chromatograms of flavonol glycosides in sea buckthorn berries (Figure S1), correlations

441

between the temperature sum variables and the harvest date of berries (Figure S2) and


20

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442

compositional response to the harvest date of berries (Figure S3) are available free of charge on

443

the ACS Publications website.

444


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References

446 447 448

1. Yang, Y. Q.; Yao, Y.; Xu, G.; Li, C. Y. Growth and physiological responses to drought and elevated ultraviolet-B in two contrasting populations of Hippophaë rhamnoides. Physiol. Plantarum 2005, 124, 431–440.

449 450

2. Yang, B. R.; Kallio, H. Composition and physiological effects of sea buckthorn (Hippophaë) lipids. Trends Food Sci. Technol. 2002, 1Hippophaë3, 160–167.

451 452 453

3. Yang, B.; Halttunen, T.; Raimo, O.; Price, K.; Kallio, H. Flavonol glycosides in wild and cultivated berries of three major subspecies of Hippophaë rhamnoides and changes during harvesting period. Food Chem. 2009, 115, 657–664.

454 455 456

4. Pop, R. M.; Socaciu, C.; Pintea, A.; Buzoianu, A. D.; Sanders, M. G.; Gruppen, H.; Vincken, J. UHPLC/PDA-ESI/MS analysis of the main berry and leaf flavonol glycosides from different carpathian Hippophaë rhamnoides L. varieties. Phytochem. Anal. 2013, 24, 484–492.

457 458 459

5. Chen, C.; Zhang, H.; Xiao, W.; Yong, Z.; Bai, N. High-performance liquid chromatographic fingerprint analysis for different origins of sea buckthorn berries. Journal of Chromatography a 2007, 1154, 250–259.

460 461

6. Belitz, H. D.; Grosch, W.; Schieberle, P. Food Chemistry, 3rd ed.; Springer Science & Business Media: Berlin, Germany, 2004; pp 806–860.

462 463 464

7. Schieber, A.; Keller, P.; Streker, P.; Klaiber, I.; Carle, R. Detection of isorhamnetin glycosides in extracts of apples (Malus domestica cv. "Brettacher") by HPLC-PDA and HPLC-APCIMS/MS. Phytochem. Anal. 2002, 13, 87–94.

465 466 467

8. Lee, J.; Mitchell, A. E. Quercetin and isorhamnetin glycosides in onion (Allium cepa L.): Varietal comparison, physical distribution, coproduct evaluation, and long-term storage stability. J. Agric. Food Chem. 2011, 59, 857–863.

468 469 470

9. Carvalho, E.; Franceschi, P.; Feller, A.; Palmieri, L.; Wehrens, R.; Martens, S. A targeted metabolomics approach to understand differences in flavonoid biosynthesis in red and yellow raspberries. Plant Physiology and Biochemistry 2013, 72, 79–86.

471 472 473 474

10. Jimenez-Aspee, F.; Quispe, C.; Soriano, M. D. P. C.; Gonzalez, J. F.; Haneke, E.; Theoduloz, C.; Schmeda-Hirschmann, G. Antioxidant activity and characterization of constituents in copao fruits (Eulychnia acida Phil., Cactaceae) by HPLC-DAD-MS/MSn. Food Res. Int. 2014, 62, 286–298.

475 476

11. Yang, L.; Ding, L.; Dou, W.; Wang, Z.; Zhang, J. New application of isorhamnetin. Patent no: CN201210187101.5.


22

Page 23 of 36


477 478 479

12. Ku, S.; Kim, T. H.; Lee, S.; Kim, S. M.; Bae, J. Antithrombotic and profibrinolytic activities of isorhamnetin-3-O-galactoside and hyperoside. Food and Chemical Toxicology 2013, 53, 197– 204.

480 481 482

13. Yang, J. H.; Shin, B. Y.; Han, J. Y.; Kim, M. G.; Wi, J. E.; Kim, Y. W.; Cho, I. J.; Kim, S. C.; Shin, S. M.; Ki, S. H. Isorhamnetin protects against oxidative stress by activating Nrf2 and inducing the expression of its target genes. Toxicol. Appl. Pharmacol. 2014, 274, 293–301.

483 484 485

14. Yang, J. H.; Kim, S. C.; Shin, B. Y.; Jin, S. H.; Jo, M. J.; Jegal, K. H.; Kim, Y. W.; Lee, J. R.; Ku, S. K.; Cho, I. J.; Ki, S. H. O-methylated flavonol isorhamnetin prevents acute inflammation through blocking of NF-κB activation. Food and Chemical Toxicology 2013, 59, 362–372.

486 487 488 489

15. Antunes-Ricardo, M.; Moreno-Garcia, B. E.; Gutierrez-Uribe, J. A.; Araiz-Hernandez, D.; Alvarez, M. M.; Serna-Saldivar, S. O. Induction of apoptosis in colon cancer cells treated with isorhamnetin glycosides from Opuntia Ficus-indica pads. Plant Foods for Human Nutrition 2014, 69, 331–336.

490 491 492 493

16. Zheng, J.; Yang, B.; Ruusunen, V.; Laaksonen, O.; Tahvonen, R.; Hellsten, J.; Kallio, H. Compositional differences of phenolic compounds between black currant (Ribes nigrum L.) cultivars and their response to latitude and weather conditions. J. Agric. Food Chem. 2012, 60, 6581–6593.

494 495 496

17. Anttonen, M. J.; Karjalainen, R. O. High-performance liquid chromatography analysis of black currant (Ribes nigrumL.) fruit phenolics grown either conventionally or organically. J. Agric. Food Chem. 2006, 54, 7530–7538.

497 498 499

18. Zheng, J.; Yang, B.; Tuomasjukka, S.; Ou, S.; Kallio, H. Effects of latitude and weather conditions on contents of sugars, fruit acids, and ascorbic acid in black currant (Ribes nigrumL.) juice. J. Agric. Food Chem. 2009, 57, 2977–2987.

500 501 502

19. Yang, B.; Zheng, J.; Laaksonen, O.; Tahvonen, R.; Kallio, H. Effects of latitude and weather conditions on phenolic compounds in currant (Ribes spp.) cultivars. J. Agric. Food Chem. 2013, 61, 3517–3532.

503 504

20. Zheng, J.; Kallio, H.; Yang, B. Effects of latitude and weather conditions on sugars, fruit acids and ascorbic acid in currant (Ribes sp.) cultivars. J. Sci. Food Agric. 2009, 89, 2011–2023.

505 506 507

21. Yang, B.; Zheng, J.; Kallio, H. Influence of origin, harvesting time and weather conditions on content of inositols and methylinositols in sea buckthorn (Hippophaë rhamnoides) berries. Food Chem. 2011, 125, 388–396.

508 509 510

22. Zheng, J.; Kallio, H.; Linderborg, K.; Yang, B. Sugars, sugar alcohols, fruit acids, and ascorbic acid in wild Chinese sea buckthorn (Hippophaë rhamnoides ssp. sinensis) with special reference to influence of latitude and altitude. Food Res. Intern. 2011, 44, 2018–2026.


23


Page 24 of 36

511 512

23. Taiz, L.; Zeiger, E. Plant Physiology, 4th ed.; Sinauer Associates: Sunderland, MA, 2006; pp. 197–344.

513 514 515

24. Ma, X.; Laaksonen, O.; Zheng, J.; Yang, W.; Trépanier, M.; Kallio, H.; Yang, B. Flavonol glycosides in berries of two major subspecies of sea buckthorn (Hippophaë rhamnoides L.) and influence of growth sites. Food Chem. 2016, 200, 189–198.

516 517 518

25. Rösch, D.; Krumbein, A.; Mügge, C.; Kroh, L. W. Structural investigations of flavonol glycosides from sea buckthorn (Hippophaë rhamnoides) pomace by NMR spectroscopy and HPLC-ESI-MSn. J. Agric. Food Chem. 2004, 52, 4039–4046.

519 520 521

26. Chen, C.; Xu, X.; Chen, Y.; Yu, M.; Wen, F.; Zhang, H. Identification, quantification and antioxidant activity of acylated flavonol glycosides from sea buckthorn (Hippophaë rhamnoides ssp sinensis). Food Chem. 2013, 141, 1573–1579.

522 523 524

27. Fang, R.; Veitch, N. C.; Kite, G. C.; Porter, E. A.; Simmonds, M. S. J. Enhanced profiling of flavonol glycosides in the fruits of sea buckthorn (Hippophaë rhamnoides). J. Agric. Food Chem. 2013, 61, 3868–3875.

525 526

28. Liu, P.; Kallio, H.; Yang, B. Phenolic Compounds in Hawthorn (Crataegus grayana) Fruits and Leaves and Changes during Fruit Ripening. J. Agric. Food Chem. 2011, 59, 11141–11149.

527 528 529

29. Halbwirth, H.; Puhl, I.; Haas, U.; Jezik, K.; Treutter, D.; Stich, K. Two-phase flavonoid formation in developing strawberry (Fragaria x ananassa) fruit. J. Agric. Food Chem. 2006, 54, 1479–1485.

530 531 532

30. Jaakola, L.; Maatta, K.; Pirttila, A. M.; Torronen, R.; Karenlampi, S.; Hohtola, A. Expression of genes involved in anthocyanin biosynthesis in relation to anthocyanin, proanthocyanidin, and flavonol levels during bilberry fruit development. Plant Physiol. 2002, 130, 729–739.

533 534 535

31. Maciej, J.; Schaeff, C. T.; Kanitz, E.; Tuchscherer, A.; Bruckmaier, R. M.; Wolffram, S.; Hammon, H. M. Bioavailability of the flavonol quercetin in neonatal calves after oral administration of quercetin aglycone or rutin. J. Dairy Sci. 2015, 98, 3906–3917.

536 537 538

32. Zhang, Y.; Wang, D.; Yang, L.; Zhou, D.; Zhang, J. Purification and characterization of flavonoids from the leaves of zanthoxylum bungeanum and correlation between their structure and antioxidant activity. Plos One 2014, 9, e105725.

539 540 541

33. Schulz, E.; Tohge, T.; Zuther, E.; Fernie, A.R.; Hincha, D.K. Natural variation in flavonol and anthocyanin metabolism during cold acclimation in Arabidopsis thaliana accessions. Plant Cell and Environment 2015, 38, 1658-1672.

542 543 544 545

34. Zheng, J.; Yang, B.; Trépanier, M.; Kallio, H. Effects of genotype, latitude and weather conditions on the composition of sugars, sugar alcohols, fruit acids and ascorbic acid in sea buckthorn (Hippophaë rhamnoides ssp. mongolica) berry juice. J. Agric. Food Chem. 2012, 60, 3180–3189.


24

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546 547

35. Wang, S. Y.; Zheng, W. Effect of plant growth temperature on antioxidant capacity in strawberry. J. Agric. Food Chem. 2001, 49, 4977–4982.

548 549 550

36. Pereira, G. E.; Gaudillere, J.; Pieri, P.; Hilbert, G.; Maucourt, M.; Deborde, C.; Moing, A.; Rolin, D. Microclimate influence on mineral and metabolic profiles of grape berries. J. Agric. Food Chem. 2006, 54, 6765–6775.

551 552


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Figure 1. Score and loading plots of PLS-DA model for sea buckthorn samples (one for each

554

variety) classified according to growth locations based on the composition of berries.

555

Abbreviations of compounds refer to Table 2.

556

Figure 2. PCA plots of the correlations between the compositional parameters of sea buckthorn

557

berries and weather variables. Abbreviations of weather variables and compounds refer to Table

558

1 and Table 2, respectively.

559

Figure 3. Correlations between the contents of flavonol glycosides in berries of sea buckthorn

560

varieties 'Terhi' and 'Tytti' and selected weather variables: (A) the sum of the daily mean

561

temperatures that are 5 °C or higher from the start of growth season until the day of harvest, (B)

562

the sum of the daily mean temperatures that are 5 °C or higher during the growth season, (C) the

563

sum of the daily mean temperatures that are 5 °C or higher in the last month before harvest, (D)

564

the average temperature in February, and (E) various variables of radiation, humidity and

565

precipitation. The correlations were studied based on samples from Sammlmäki, samples from

566

Kittilä, and samples from both Sammalmäki and Kittilä. Abbreviations of weather variables and

567

compounds refer to Table 1 and Table 2, respectively.


26

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Table 1. Weather Variables and Their Abbreviations abbreviations Dgs SUMTgs SUMTgh SUMTm HDgh HDm Tm Tw TDm MinTm LTm MaxTm HTm Tjan...Tsep MaxTjan*MaxTsep MinTjan*MinTsep Rgh Rm Rw Rjan... Rsep Pgh Pm Pw Pjan... Psep Hgh Hm Hw Hjan...Hsep DH20to30gh DH30to40gh DH40to50gh DH50to60gh DH60to70gh DH70to80gh DH80to90gh DH90to100gh DH50to60m DH60to70m DH70to80m DH80to90m DH90to100m

weather variables growth season period (day) sum of the daily mean temperatures that are 5 °C or higher during the growth season (°C) sum of the daily mean temperatures that are 5 °C or higher from the start of growth season until the day of harvest (°C) sum of the daily mean temperatures that are 5 °C or higher in the last month before harvest (°C) hot days (temperature > 25 °C ) from the start of growth season until the day of harvest (day) hot days (temperature > 25 °C ) in the last month before harvest (day) average temperature in the last month before harvest (°C) average temperature in the last week before harvest (°C) mean daily temperature difference in the last month before harvest (°C) minimum temperature in the last month before harvest (°C) average of daily lowest temperature in the last month before harvest (°C) maximum temperature in the last month before harvest (°C) average of daily highest temperature in the last month before harvest (°C) average temperature in January...September (°C) maximum temperature in January...September (°C) minimum temperature in January...September (°C) radiation from the start of growth season until the day of harvest (kJ/m2) radiation during the last month before harvest (kJ/m2) radiation during the last week before harvest (kJ/m2) radiation in January...September (kJ/m2) precipitation from the start of growth season until the day of harvest (mm) precipitation in the last month before harvest (mm) precipitation in the last week before harvest (mm) precipitation in January...September (mm) average humidity from the start of growth season until the day of harvest (%) average humidity in the last month before harvest (%) average humidity in the last week before harvest (%) average humidity in January...September (%) percentage of the days with relative humidity 20-30% from the start of growth season until the day of harvest (%) percentage of the days with relative humidity 30-40% from the start of growth season until the day of harvest (%) percentage of the days with relative humidity 40-50% from the start of growth season until the day of harvest (%) percentage of the days with relative humidity 50-60% from the start of growth season until the day of harvest (%) percentage of the days with relative humidity 60-70% from the start of growth season until the day of harvest (%) percentage of the days with relative humidity 70-80% from the start of growth season until the day of harvest (%) percentage of the days with relative humidity 80-90% from the start of growth season until the day of harvest (%) percentage of the days with relative humidity 90-100% from the start of growth season until the day of harvest (%) percentage of the days with relative humidity 50-60% in the last month before harvest (%) percentage of the days with relative humidity 60-70% in the last month before harvest (%) percentage of the days with relative humidity 70-80% in the last month before harvest (%) percentage of the days with relative humidity 80-90% in the last month before harvest (%) percentage of the days with relative humidity 90-100% in the last month before harvest (%)


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Table 2. Flavonol Contents (Micrograms per 100 g of Fresh Berries) in Sea Buckthorn Berries (H. rhamnoides ssp. rhamnoides)a. isorhamnetin-3-O sophoroside-7-O rhamnoside b

isorhamnetin-3-O isorhamnetin-3-O glucoside-7-O isorhamnetin-3- isorhamnetin-3- rutinoside-7-O rhamnoside O -rutinoside O -glucoside glucoside

isorhamnetinglucosiderhamnoside 1



isorhamnetinglucoside

total unknown isorhamnetinglycosides

(I-3-S-7-Rh)

(I-3-G-7-Rh)

(I-3-R)

(I-3-G)

(I-3-R-7-G)

(I-G-Rh 1)

(I-G-Rh 3)

(I-G-Rh 4)

(I-G)

(Unk I-gly)

comparison between varieties Terhi (n = 208) Tytti (n = 208)

7.41±1.83 a 8.92±3.03 b

25.67±10.45 a 26.38±10.63 a

30.40±6.85 b 18.27±4.66 a

9.31±3.44 a 12.42±6.08 b

1.78±0.63 a 2.64±0.50 b

5.90±1.84 a 8.98±3.37 b

1.38±1.81 a 3.59±0.96 b

1.67±1.11 a 2.52±1.31 b

1.98±0.35 a 3.05±0.95 b

6.32±1.19 a 8.64±3.13 b

comparison between locations Terhi S (n = 112) 2007-2013 K (n = 96) 2009-2013

5.96±0.67 E 9.10±1.19 F

17.81±5.03 E

26.35±4.91 E

11.14±3.39 F

1.75±0.36 E

4.55±0.83 E

0.89±1.21 E

2.45±0.52 F

1.96±0.36 E

35.12±5.66 F 15.65±3.86 j 21.10±3.70 k

7.17±1.94 E 16.82±5.21 k 7.66±1.99 j

1.82±0.85 E 2.59±0.38 j 2.70±0.60 j

7.47±1.39 F 6.32±1.59 j 11.85±2.22 k

1.94±2.20 F 3.19±0.49 j 4.03±1.14 k

0.76±0.91 E 3.44±0.65 k 1.52±1.08 j

2.01±0.33 E 3.67±0.84 k 2.37±0.51 j

6.35±0.96 E 6.29±1.43 E

6.36±0.69 j 11.69±1.94 k

34.84±7.12 F 17.98±5.78 j 35.45±6.33 k

11.47±1.19 k 5.58±0.90 j

13.77±0.63 17.03±1.37

65.65±4.33 68.46±7.57

60.61±5.23 37.74±5.72

8.93±1.13 10.30±1.75

3.79±0.22 4.81±0.50

13.70±1.08 24.36±3.44

trace 6.27±0.88

none none

4.06±0.23 5.23±0.77

11.10±0.38 8.96±0.70

variety location

Tytti

S (n = 108) K (n = 100)

year

2007-2013 2009-2013

flavonol glycosides in unripe berries Terhi K (n = 20) 2008 Tytti K (n = 20) 2008

quercetin-3-O sophoroside-7-O rhamnoside (Q-3-S-7-Rh) comparison between varieties Terhi (n = 208) Tytti (n = 208) comparison between locations Terhi S (n = 112) 2007-2013 K (n = 96) 2009-2013 Tytti S (n = 108) 2007-2013 K (n = 100) 2009-2013 flavonol glycosides in unripe berries Terhi K (n = 20) 2008 Tytti K (n = 20) 2008

quercetin-3-O - quercetin-3-O rutinoside glucoside (Q-3-R) (Q-3-G)

quercetin-3-O glucoside-7-O rhamnoside (Q-3-G-7-Rh)

monoglycosides of isorhamnetin (I-monogly)

di-glycosides of isorhamnetin (I-digly)

tri-glycosides of isorhamnetin (I-trigly)

di-glycosides of quercetin (Q-digly)

total quercetin glycosides (Total QG)

total isorhamnetin glycosides (Total IG)

total flavonol glycosides (Total FG)

2.45±0.55 a 3.32±0.93 b

3.56±0.94 a 3.94±1.27 b

2.72±0.50 a 4.18±1.09 b

2.69±0.87 a 3.32±1.06 b

11.24±3.67 a 15.47±6.98 b

64.63±19.04 b 59.74±18.05 a

9.15±2.00 a 11.57±3.09 b

6.22±1.63 a 7.25±2.11 b

11.43±2.24 a 14.75±3.04 b

91.83±21.25 a 95.42±18.16 a

103.25±23.03 a 110.16±20.79 b

2.01±0.15 E 2.96±0.39 F 2.55±0.21 j 4.14±0.66 k

3.20±0.50 E 4.00±1.14 F 3.30±0.99 j 4.63±1.19 k

2.76±0.52 E 2.67±0.46 E 4.56±1.14 k 3.76±0.86 j

2.05±0.47 E 3.45±0.57 F 2.52±0.72 j 4.18±0.59 k

13.10±3.67 F 9.06±2.19 E 20.49±6.00 k 10.04±2.35 j

52.05±10.33 E 79.30±16.16 F 46.58±11.80 j 73.96±11.73 k

7.71±0.79 E 10.83±1.63 F 8.95±0.88 j 14.39±1.91 k

5.24±0.86 E 7.37±1.56 F 5.81±1.42 j 8.81±1.56 k

10.01±1.39 E 13.07±1.89 F 12.93±2.61 j 16.71±2.12 k

79.22±14.14 E 106.53±18.56 F 87.50±17.74 j 103.97±14.39 k

89.23±15.28 E 119.61±19.51 F 100.42±20.27 j 120.68±15.63 k

5.60±0.21 7.46±0.87

8.31±0.55 10.17±1.54

4.43±0.43 5.96±0.80

7.46±0.82 12.65±2.45

12.99±1.32 15.53±2.46

139.96±10.47 136.83±17.31

17.57±0.64 21.84±1.80

15.77±1.27 22.82±3.11

25.79±1.73 36.25±4.33

181.62±11.89 183.16±21.98

207.41±13.37 219.41±25.85

a

Significant difference (p < 0.05) between samples of different varieties are marked as a–b, and between samples grown at different locations (each variety compared separately) as E–F and j–k for Terhi and Tytti, respectively. b S, Sammalmäki, Finland; K, Kittilä, Finland.


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Figure 1.


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Figure 2.


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Fig. 3A


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Fig. 3B


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Fig. 3C


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Fig. 3D


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Fig. 3E


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Graphic for table of contents


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Sea Buckthorn (HippophaÃ« rhamnoides ssp. rhamnoides) Berries in

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