JFS S: Sensory and Food Quality Relation between Developmental Stage, Sensory Properties, and Volatile Content of Organically and Conventionally Grown Pac Choi (Brassica rapa var. Mei Qing Choi) MARTIN TALAVERA-BIANCHI, KOUSHIK ADHIKARI, EDGAR CHAMBERS IV, EDWARD E. CAREY, AND DELORES H. CHAMBERS Practical Application: The increased popularity of organic production has amplified the need for research that will help in understanding how this production system affects the final quality of food products. This study suggests that the stage of development has a much larger impact on sensory quality than organic or conventional growing of pac choi. Findings from this study promote consumer choice by showing that comparable sensory quality can be obtained using either production system, making the ultimate choice not only based on sensory quality but consumer choice related to environmental beliefs or economics. Keywords: bok choy, organic, pac choi, sensory, volatiles Introduction O rganic foods often are promoted as being more environmentally friendly. Crop rotation, cover crops, and natural products (such as natural fertilizes and pesticides) are used to enhance or maintain long-term soil fertility, minimize pollution, avoid synthetic fertilizers and pesticides, consider the social and economic impact, and produce higher quality products (Bourn and Prescott 2002; Winter and Davis 2006). However, results at this point seem inconsistent and show no clear trends or patterns regarding the effects that organic fertilization have on the crops’ final quality (Basker 1992; Bourn and Prescott 2002). Those inconsistencies may exist because the differences between organic and conventional practices are product specific (Fillion and Arazi 2002) or because differences are dependent on confounding factors such as age, picking time, or transportation. MS 20090596 Submitted 6/25/2009, Accepted 2/2/2010. Author TalaveraBianchi is with PepsiCo Inc., Sensory & Consumer Sciences, Barrington, IL 60010, U.S.A. Authors Adhikari, Chambers IV, and Chambers are with The Sensory Analysis Center, Dept. of Human Nutrition, Justin Hall, Kansas State Univ., Manhattan, KS 66506, U.S.A. Author Carey is with Dept. of Horticulture, Forestry, and Recreation Resources, Throckmorton, Kansas State Univ., Manhattan, KS 66506, U.S.A. Direct inquiries to author Chambers (E-mail: delores@ksu.edu). R 2010 Institute of Food Technologists doi: 10.1111/j.1750-3841.2010.01585.x  C Further reproduction without permission is prohibited Pac choi (Brassica rapa var. Mei Qing Choi) is a variety of Chinese cabbage, well known in Asia, that is gaining popularity in the United States. The flavor of pac choi has been previously studied by Schnitzler and Kallabis-Rippel (1998). Those authors studied different varieties of cooked and fresh pac choi using a trained sensory panel. Descriptive terms used by these authors were sweet, sour, bitter, spicy, and cabbage-like. Other studies focused on instrumental analysis of pac choi leaves to evaluate flavonoid composition (Rochfort and others 2006), phenolic content in organic plants (Young and others 2005), and the effect of packaging on their shelf life (Lu 2007). However, stage of development was not included in these studies. Stage of development at the time of harvest should be included when evaluating pac choi because this plant is frequently consumed at different maturity levels (that is, baby or mature stage) (Rochfort and others 2006). Plant maturity at the time of harvest is critical for flavor and texture development (Mattheis and Fellman 1999). It has been suggested that age has an effect on the content of flavor compounds such as catechins and amino acids, which tend to create off-flavors in young tea leaves (Kinugasa and others 1997). It also has been suggested that some fruits such as muskmelon must be harvested at their ripening stage for best postharvest quality (Asghary and others 2005). Tomatoes, where harvest maturity is a critical factor related to sensory properties, have been the object of many studies to Vol. 75, Nr. 4, 2010—JOURNAL OF FOOD SCIENCE S173 S: Sensory & Food Quality ABSTRACT: This study was conducted to identify and quantify the sensory characteristics and chemical profile of organically and conventionally grown pac choi (Brassica rapa var. Mei Qing Choi), also called bok choy, at 3 stages of growth (2.5, 4.5, and 6.5 wk). Sensory and instrumental data were correlated using partial least squares regression. Pac choi was grown in late spring. Descriptive sensory analysis was conducted by a highly trained panel and compounds were identified and quantified using a gas chromatograph/mass spectrometer. The findings of the study indicate that the differences in sensory characteristics and chemical profiles among stages of growth are more substantial than the differences between organic and conventional production. Green-unripe, musty/earthy, lettuce, and sweet flavors are representative in pac choi at early stages of growth. When older, pac choi has higher intensities of green-grassy/leafy, bitter, cabbage, and sulfur flavors that are associated with the increase of (Z)-3hexen-1-ol, octyl acetate, 1-nonanol, 2-decanone, 1-penten-3-ol, linalool, camphor, menthol, isobornyl acetate, geranylacetone, and cedrol compounds. Conventional pac choi was higher than organic pac choi in green overall, bitter, and soapy flavors only at 2.5 wk of age. This may be associated with the presence of (Z)-3-hexenal, 2-hexyn-1-ol, and (E)-2-hexenal compounds. Maturity stages of pac choi . . . assess the relation between fruit ripeness, sensory properties, and chemical composition (Hayase and others 1984; Shewfelt and others 1988; Stern and others 1994; Yilmaz and others 2002). The relation between sensory properties and chemical composition has received important attention in the past. Studies that relate sensory and instrumental data have been conducted for products such as wheat bread (Quı́lez and others 2006), virgin olive oil (Morales and others 1995), durian fruit (Voon and others 2007), navel oranges (Baxter and others 2005), and exotic salad crops (Price and others 1990), using different statistical methodologies. A study focused on wine flavor (Noble and Ebeler 2002) explored and compared 3 different multivariate methodologies that can be used to relate sensory and instrumental data. These studies used principal component analysis, generalized procrustes analysis, and partial least squares regression. Authors concluded that all 3 methods provide similar results. However, some differences were noted. Many studies have been able to link aroma volatiles found in foods with sensory characteristics. For example, butyl acetate, 1-hexanal, and camphor are aroma volatiles found in apples linked to fruity, green, and piney odors respectively (Mehinagic and others 2006). Pentanal, heptanal, and octanal are related to nutty, floral, and citrus aromas in rice (Yang and others 2008). Similarly, hexanal, nonanal, and acetaldehyde are related to green, soapy, and fruity aromatics in grapefruit juice (Buettner and Schieberle 2001). Bott and Chambers (2006) found combinations of chemicals that produced beany odors and Hongsoongnern and Chambers (2008a) related chemicals to “green” characteristics found in various food products. The objectives of this study are (1) to evaluate the sensory characteristics and the aroma volatile content of organically and conventionally grown pac choi leaves at 3 stages of development and (2) to correlate sensory and instrumental data by means of partial least squares regression. S: Sensory & Food Quality Materials and Methods border of each plot. Compost application rates were based on the assumption that 50% of the nitrogen from compost would be available to plants during the growing season, whereas 100% would be available from conventional fertilizers (Warman and Havard 1997). Low- and high-fertility plots were fertilized with equal amounts of compost or synthetic fertilizer at the beginning of the growing season, and high-fertility plots received additional fertilization during the growing season as described later. Pac choi (Brassica rapa L. chinensis “Mei Qing Choi”) (Johnny’s Selected Seed, Albion, Maine, U.S.A.) and tomato (Lycopersicon esculentum “Bush Celebrity”) (Totally Tomatoes, Randolph, Wis., U.S.A.) were grown in one half of each open field or high tunnel plot (6.8 m × 3 m) in 2007 and 2008, with a rotation between pac choi and tomato plots each year. In our experimental system, a spring and a fall crop of pac choi was grown each year, whereas a single crop of tomato was grown. Between the spring and fall pac choi crops, plots were seeded with a summer cover crop of buckwheat (Fagopyrum sagittatum) (Albert Lea Seed, Albert Lea, Minn., U.S.A.) at a rate of 134 kg/ha. In the late fall, all plots were seeded with a cover crop of annual rye (Secale cereale) (Albert Lea Seed, Albert Lea, Minn., U.S.A.) at a rate of 229 kg/ha. Conventional high- and low-fertility plots were fertilized with Jack’s Professional Peat-lite N–P2 O5 –K2 O 20–10–20 (Allentown, Pa., U.S.A.) at a rate of 98 kg/ha. Organic plots received MicroLeverage compost N–P2 O5 –K2 O 0.6–0.8–0.5 (Hughesville, Mo., U.S.A.) at a rate of 197 kg/ha. Only pac choi grown in the outside plots with low amounts of fertilizer were used for this specific study. Pac choi transplants were started in a greenhouse in Sunshine Mix Special Blend E6340 (SunGro Horticulture, Bellevue, Wash., U.S.A.) supplemented with MicroLeverge compost. Pac choi was planted on April 1, 2008 and harvested on April 20 (2.5 wk old for baby pac choi), May 5 (4.5 wk old for optimum growth), and May 19 (6.5 wk old for overgrown pac choi). Testing times were selected based on the assumption that pac choi is at an optimal growth level at 1 mo after planting. Samples Sample preparation Trials were conducted at the K-State Horticulture Research and Extension Center, Olathe, Kans., U.S.A., on experimental plots established in 2002 for comparison of crops grown under organic and conventional production systems in high tunnels (unheated, passively ventilated greenhouses) and open field plots (Zhao and others 2007). The soil was a Kennebec silt loam. Six 9.8 m × 6.1 m high tunnels with 1.5 m sidewalls (Stuppy, North Kansas City, Mo., U.S.A.) and 6 adjacent 9.8 m × 6.1 m field plots were used for this study. High tunnels were covered with single layer 6-mil (0.153 mm) K-50 polyethylene (Klerk’s Plastic Product Manufacturing Inc., Richburg, S.C., U.S.A.). At establishment of the experimental plots, the 6 high tunnels were divided into 3 groups (blocks) and the 2 high tunnels in each block were randomly assigned for longterm conventional or organic management treatments. A similar setup was used in the field plots. Organic plots were managed in compliance with USDA Natl. Organic Program standards, and were inspected and certified in 2003, 2006, 2007, and 2008. For this study, beginning in 2007, each high tunnel or openfield plot was subdivided into three 3.2 m × 6.1 m plots to which 1 of 3 fertilizer levels were assigned (high, low, and no fertilizer), following a Latin square design to avoid bias due to position effects in the high tunnels. Fertilizer rates were determined based on soil analysis at the beginning of the study in 2007, and recommendations for vegetable crops in Kansas (Marr and others 1998), with compost applied to organic plots and synthetic fertilizer applied to conventional plots. A buffer zone was included in the Sensory analysis. Plants were harvested 1 to 3 d before testing. After harvest, the plants were immediately rinsed using cold tap water to remove excess dirt. To minimize postharvest effects, plants were stored in a refrigerated container for transport to the Kansas State Univ. campus located in Manhattan, Kans., U.S.A., and moved into a walk-in refrigerator for storage at 4 ◦ C immediately after arrival. Samples remained in the refrigerator until testing. The plants were sprayed daily with tap water to maintain moisture. On the day of testing, plants were retrieved from the refrigerator. Random leaves of similar visual characteristics were removed from each stalk (not including the stem) and rinsed using distilled water. Excess water was eliminated with a salad spinner (Oxo Intl. Ltd., New York, N.Y., U.S.A.). Samples were served to the panelists monadically in 6 in. foam plates identified with a 3-digit code to eliminate potential panelist bias. The sample amount was dependant on leaf size. For example, for baby pac choi (2.5-wk-old plant) 1 whole sprig composed of several leaves was served to each panelist, 1 to 2 leaves were served to each panelist when leaves were 4.5 and 6.5 wk old. Volatile analysis. The same day of sensory testing, approximately 5 to 10 g from leaves of each treatment were vacuum sealed and frozen at −80 ◦ C for 30 d until volatile analysis. The day of the analysis, samples were retrieved from the freezer and thawed at room temperature (22 ± 1 ◦ C) for approximately 30 min. For solidphase microextraction (SPME) sampling, 4 g pac choi leaves were blended with 200 mL of reverse osmosis, deionized, carbon-filtered S174 JOURNAL OF FOOD SCIENCE—Vol. 75, Nr. 4, 2010 Maturity stages of pac choi . . . Panelists Six highly trained panelists from the Sensory Analysis Center at Kansas State Univ. (Manhattan, Kans., U.S.A.) were used for this study. The trained panel was formed by 5 females and 1 male with ages ranging from 45 to 65 y old. The panelists had completed more than 120 h of descriptive training, averaged more than 2000 h of testing experience, and had prior experience testing vegetables and vegetable products. Panelist performance is evaluated during orientation/training by examining individual daily results of samples presented to the panelists. This is done both by the panel leader in the panel session and by the project manager after the session. Additional training is provided if panelists need practice for consistent evaluation. Experimental procedure Sensory analysis. This lexicon for pac choi was previously developed by Talavera-Bianchi and others (2010) to describe flavor of different leafy vegetables and contains terms, references, and reference preparation techniques. A lexicon consisting of 29 terms with definitions and references was presented to the panelists along orientation samples in one 90-min orientation session prior the start of testing so they could become familiar with the terminology, test procedures, and samples. The original lexicon consisted of 26 flavor and mouth feel attributes. However, 3 texture attributes were added because the authors believed that this would aid in describing changes in the plant during the maturation process. Similar lexicons have been developed and used for other products such as green tea (Lee and Chambers 2007), tomatoes (Hongsoongnern and Chambers 2008b), ice cream (Thompson and others 2009), and brewed coffee (Seo and others 2009). The day of testing, panelists were presented with the lexicon and references used during orientation. Data were collected using a computerized collection system (Compusense Five version 4.4.8, 2002, Guelph, Ontario, Canada). Intensities for each attribute were recorded using a 0 to 15 point scale divided in 0.5-point increments, 0 meaning “none” and 15 meaning “extremely high.” Panelists evaluated the samples individually and followed a completely randomized block design with the stage of development as the blocking factor. A total of 3 d of testing were conducted following each individual harvest date. Six samples of pac choi were evaluated in each of three 90-min sessions. Panelists concentrated on the leaf portion only. Reverse osmosis, deionized, carbon-filtered water, and unsalted crackers were used to rinse the palate between the samples. A similar procedure has been used in the past to evaluate the sensory characteristics of 4 samples of calcium-biofortified lettuce (Park and others 2009). Gas chromatography–mass spectrometry. Volatile compounds were identified and quantified using a Varian Saturn CP-3800 Gas Chromatograph/Mass Spectrometer 2200 (Varian Inc., Walnut Creek, Calif., U.S.A.). The sample vials were equilibrated at 40 ◦ C/500 rpm for 10 min. SPME was performed using a StableFlex Divinylbenzene/Carboxen/Polydimethylsiloxane 50/30 µm fiber (Sigma Aldrich, St. Louis, Mo., U.S.A.) for 20 min at 40 ◦ C. The agitation during extraction was of 250 rpm. The extracted compounds were thermally desorbed at 250 ◦ C for 3 min in the front injection port of the gas chromatograph. After  R the injection, the fiber was baked at 270 ◦ C for 30 min. An RTX -5 Capillary Column (30 m length × 0.25 mm internal dia. × 0.25 µm film thickness; Restek U.S., Bellefonte, Pa., U.S.A.) was used to separate the volatiles desorbed from the fiber. The initial temperature of the column was set at 40 ◦ C for 2 min and then raised to 200 ◦ C at a rate of 5 ◦ C/min−1 and held for 1 min (total GC run time was 35 min). This process was optimized by conducting several practice runs prior to the beginning of this study. Varian MS Workstation software (version 6.8) was used for system control, data collection, and data processing. Compound identification was based on NIST 2005 version 2.0 Mass Spectra library search. The compounds’ final concentration was based on the concentration of the internal standard, which had a known amount. Three replications were analyzed for each treatment. Kovats retention indices were calculated to aid in the identification of the volatile compounds. A blend of hydrocarbon (HC) mix and carbon disulfide (1 drop of HC mix in 1 mL of CS2 directly injected to the GC) was also run under the same methodology to generate the retention times of the n-alkanes (C6 –C20 ) for calculating the Kovats indices. Comparing Kovats indices from chemicals previously identified using the same column and stationary phase under similar conditions has shown to be an accurate method of identification (Moustafa 2008). Analysis Analysis of variance (ANOVA) with PROC MIXED (panelist and replication as the random effects) was used to detect overall differences among treatments for individual sensory attributes. PROC GLM (3 replications) was used to detect differences for individual  R volatile compounds. ANOVA was computed in SAS (2002, version 9.1.3; SAS Inst., Cary, N.C., U.S.A.). Partial least squares regression (PLS2) was used to correlate sensory and instrumental data. PLS is a soft-modeling method, which is widely used to predict a set of dependant variables (sensory attributes) from a set of independent variables (volatile compounds) (Noble and Ebeler 2002). This method is particularly useful when there is a need to predict a set of dependant variables from a very large set of independent variables, which is the case in this study (Abdi 2003). PLS has been previously used to correlate instrumental and sensory data in cheese (Hough and others 1996), diced tomatoes (Lee and others 1999), and ice cream (Chung and others 2003). Even though this analysis does not determine which volatile components are actually responsible for specific sensory attributes, it does help in studying the relationship between certain volatiles and sensory characteristics (Noble and Ebeler 2002). This analysis was performed using Unscrambler (2005, version 9.2; Camo Process AS, Oslo, Norway). Results and Discussion Sensory analysis Findings from the study show that stage of development is an important factor affecting sensory characteristics of pac choi. Twenty-one flavor and texture attributes were significantly different among maturity levels (P-value ≤ 0.05) (Table 1). Most of the attributes’ intensities increased as the plants get older, although some decreased. For example, attributes such as crispness, fiber awareness, overall green, green-grassy-leafy, woody, sulfur, soapy, toothetch, and bitter had lower intensities in younger plants and higher intensities in older plants. The attributes that remained stable throughout the plant development process were green-viney, radish, water-like, petroleum-like, pungent, bite, and the sour taste. Vol. 75, Nr. 4, 2010—JOURNAL OF FOOD SCIENCE S175 S: Sensory & Food Quality water using an electric hand blender (Rival, Peoria, Ill., U.S.A.) for 20 s. The mixture was then filtered through double-layered cheese cloth. From the filtered solution, 1 mL was transferred to a 10 mL clear headspace vial and mixed with 0.2 g of sodium chloride (NaCl). In addition, 5 µL of 0.2 ppm 1,3 dichlorobenzene in methanol (internal standard) was added. The internal standard was selected based on previous trials. Glass vials were closed using an open-center screw cap with a 1.8 mm silicone/PTFE septum (Varian, Palo Alto, Calif., U.S.A.). Maturity stages of pac choi . . . The typical green, bitter, and sulfur flavors of pac choi and other vegetables of the Brassica family are believed to be caused by glucosinolate-derived compounds. The main glucosinolates found in pac choi are 3-butenyl- and 1-methoxy-3-indoylmethyl (He and others 2003). It would be expected that as the concentration of S: Sensory & Food Quality these compounds increase when the plant matures, the intensity of typical flavors may increase as well. When glucosinolates are released from the plant cells, they are broken down by enzymatic action into products such as nitriles and isothiocyanates, which are also responsible for the “hot and spicy” flavors of mustard, radishes, and other plants from the Brassica family (Johnson 2001). The lower concentration of glucosinolates in pac choi compared to Table 1 --- Analysis of variance showing the significant differences (95% confidence) between stages of devel- other plants of the Brassica family is consistent with its milder flaopment for individual attributes for both organic and vor (He and others 2003). Contrarily, attributes such as moistness, conventional systems. green-unripe, and overall sweet had higher intensity in younger plants and lower intensities in older plants. It may be that as the Stage of development plant matures, sugar may be used by the plant and the development Attributesa,b Fertilization 2.5 wk 4.5 wk 6.5 wk of other flavor characteristics such as sulfur or bitterness may mask Crispness Organic 2.7b 3.2ab 3.5a the sweet taste as well as reducing the perception of unripeness in Conventional 2.7b 3.3a 3.6a pac choi. Many of these flavor characteristics have been reported Moistness Organic 5.5a 4.1c 4.5b for pac choi in the past. Schnitzler and Kallabis-Rippel (1998) used Conventional 5.7a 4.2c 4.6b terms such as sweet, sour, bitter, and spicy to describe flavor of Fiber awareness Organic 2.9c 4.3b 4.7a Conventional 3.3c 4.2b 4.9a raw pac choi. In our study, we also used the sour and spicy (bite) Overall green Organic 5.9b 5.9b 7.0a attributes. However, they were not significantly different among Conventional 6.9a 6.1b 7.0a stages of maturity or production system (that is, organic and conGreen-unripe Organic 1.6a 1.0b 0.9b ventional). Conventional 1.7a 0.9b 0.9b Green-peapod Organic 1.3a 0.4b 1.3a Few differences were found between organically and convenConventional 1.5a 0.4b 1.1a tionally grown pac choi. The few small differences that exist were Green-grassy/leafy Organic 4.6b 5.0b 6.0a found only at the 2.5-wk stage of development (Table 2). In this Conventional 5.3ab 5.0b 5.8a case, conventionally grown pac choi had significantly higher intenGreen-viney Organic 1.6 1.8 1.9 sities (P-value ≤ 0.05) of overall green, soapy, bitter, and petroleumConventional 1.6 2.0 1.8 Cabbage Organic 2.4 2.6 2.7 like attributes. No differences were found at 4.5- or 6.5-wk-old pac Conventional 2.2b 2.6ab 2.8a choi. This suggests that the effect of organic production may be Lettuce Organic 1.8a 1.9a 1.4b more evident at early stages of development. It has been suggested Conventional 1.7 1.8 1.5 that organic treatment may increase the opportunity of insect atSpinach Organic 1.6b 2.0a 1.9a Conventional 1.7b 2.0a 1.8ab tack in pac choi, which may cause the amount of total phenolics Parsley Organic 0.9b 1.4a 1.1b to increase, affecting its flavor (Young and others 2005). KobueConventional 1.1b 1.5a 1.1b Lekalake and others (2007) suggest that phenolic compounds inRadish Organic 2.0 1.9 1.8 crease the bitterness and astringency of sorghum grains. Another Conventional 1.9 1.9 2.0 study also reported higher bitterness in organically grown carrots Piney Organic 1.1a 0.4b 0.9a Conventional 1.6a 0.8b 0.8b (Haglund and others 1999). In our study, bitterness was lower in the Woody Organic 1.3b 1.5b 2.0a organic pac choi than in the conventional pac choi at 2.5 wk matuConventional 1.3b 1.7a 1.9a rity, but there were no differences in astringency. Water-like Organic 1.6 1.9 1.6 Conventional 1.6 1.8 1.7 Musty/earthy Organic 2.3 2.6 2.3 Gas chromatography–mass spectrometry Conventional 2.9a 2.3b 2.4b Forty-eight volatile compounds were identified and quantified Sulfur Organic 1.0c 1.7b 2.2a (Table 3). The chemicals that were mostly present in pac choi Conventional 1.3c 1.7b 2.3a leaves are the aldehydes (Z)-3-hexenal (9), (E)-2-hexenal (11), (E,E)Soapy Organic 0.5c 1.2b 1.6a Conventional 1.1b 1.3ab 1.5a 2,4-hexadienal (15), and benzeneacetaldehyde (21); alcohols such Petroleum-like Organic 0.4 0.3 1.0 as 2-hexyn-1-ol (10), (Z)-3-hexen-1-ol (12), (E)-3-hepten-1-ol (13), Conventional 1.2 0.5 0.7 and (E)-2-nonen-1-ol (27); as well as noncyclic and cyclic HCs Pungent Organic 2.1 2.0 2.0 such as 4,5-dimethylthiazole (30) and isothiocyanato-cyclohexane Conventional 2.2 2.1 2.1 Bite Organic 1.9 2.1 2.3 (40), respectively. Many of these compounds have been previConventional 1.9 2.2 2.2 ously reported as providing “green” aromas in foods. For example, Toothetch Organic 1.1b 2.0a 2.3a Conventional 1.3b 2.1a 2.1a Overall sweet Organic 1.4a 1.3ab 1.1b Table 2 --- Individual attributes that showed significant difConventional 1.3ab 1.5a 1.2b ferences (P -value ≤ 0.05) between organic and convenSour Organic 1.6 1.5 1.6 tional pac choi at the baby stage (2.5 wk).a Conventional 1.6 1.6 1.6 Bitter Organic 4.9b 7.0a 7.0a Attributes Organic Conventional Conventional 6.3 6.6 6.7 b Overall green 5.9b 6.9a Salty Organic 0.6ab 0.4b 0.9a Soapyb 0.5b 1.1a Conventional 0.5b 0.6ab 0.9a Bitterb 4.9b 6.3a Umami Organic 2.1a 2.1a 1.7b Petroleum-likec 0.4b 1.2a Conventional 2.0 2.1 1.9 a Astringent Organic 1.4b 1.8a 1.9a No differences between organic and conventional pac choi were observed at 4.5 or 6.5 wk maturity. Conventional 1.8 1.8 1.9 b a Significant differences are at the 95% confidence level. Different letters indicate statistically significant differences. b Different letters (a, b, c) show differences among the 3 stages of development. S176 JOURNAL OF FOOD SCIENCE—Vol. 75, Nr. 4, 2010 Significant differences are at the 95% confidence level. Different letters indicate statistically significant differences. c Significant differences are at the 90% confidence level. Different letters indicate statistically significant differences. Maturity stages of pac choi . . . Table 3 --- Retention time, retention index, and concentration (pg per g) of 48 volatile aromatics found in organically and conventionally grown pac choi at 3 stages of development. 1 2-butanone 2 Methyl propionate 3 3-methyl-2-butanone 4 1-penten-3-ol Literature aroma Fragrant pleasant (Morales and others 1995) Grassy green (Chida and others 2004) 5 Butanoic acid, methyl ester 6 2-methyl-3-pentanone 7 (E)-2-hepten-1-ol 8 2-methyl-butanoic acid, methyl ester 9 (Z)-3-hexenal Green grassy (Morales and others 1995; Iraqi and others 2005) 10 2-hexyn-1-ol 11 (E)-2-hexenal Green green-fruity bitter (Morales and others 1995; Klesk and Qian 2003) 12 (Z)-3-hexen-1-ol Green banana-like (Morales and others 1995; Chambers and others 1998) 13 (E)-3-hepten-1-ol 14 Heptanal Floral (Yang and others 2008) 15 (E,E)-2,4-hexadienal Green vegetable burnt (Hartvigsen and others 2000; Venkateshwarlu and others 2004) 16 1-isothiocyanatobutane 17 1-octen-3-ol 18 4-isothiocyanato-1Sulfur (Engel and others 2002) butene 19 Octanal Green citrus-like (Buettner and Schieberle 2001; Iraqi and others 2005) 20 (E)-2-octenal Green (Aparicio and others 1997) 21 Benzeneacetaldehyde Green flower honey bitteralmond (Heiniö and others 2003) 22 4-methyl-1-undecene 23 1-octanol Grass pepper (Mehinagic and others 2006) 24 4-ethyl-5methylthiazole 25 2-nonanone 26 3,7-dimethyl-1,6Sweet floral citrus woody (Thi octadien-3-ol Minh Tu and others 2002; (linalool) Chida and others 2004) 27 (E)-2-nonen-1-ol 28 Isopinocarveol 29 1-nonanol 30 4,5-dimethyl-thiazole Smoky roasty fragrant nutty (Specht and Baltes 1994) 31 Acetic acid, octyl ester 32 Benzyl nitrile 33 Camphor Piney spicy (Mehinagic and others 2006) 34 (Z)-6-nonenal 35 4-methylpentyl isothiocyanate 36 Menthol Refreshing light sweet pungent (Chida and others 2004) 37 2-decanone 38 (E)-2-decen-1-ol 39 2,6,6-trimethyl-1cyclohexene-1carboxaldehyde 40 Isothiocyanatocyclohexane 41 Isobornyl acetate Treatments1,2,3 RI5 O2.5 C2.5 O4.5 C4.5 O6.5 2.5 585.6 26.7 25.9 21.2 18.7 24.2 20.7 2.8 3.1 3.3 622.2 657.3 685.1 7.5 8.2 13.2b 7.9 7.6 14.8b 5.6 10.8 25.4a 5.1 8.5 21.2a 6.1 10.0 20.5a 5.5 8.9 31.8a 3.9 728.1 44.8a 34.9b 41.4a 33.8b 36.1a 31.6b 4.4 4.9 5.0 754.1 775.7 781.7 7.8a 16.8b 32.7 6.9b 19.3b 29.1 7.5a 25.2ab 32.3 5.7b 19.5ab 29.5 7.4a 22.9a 29.1 5.9b 29.8a 29.5 5.5 805.7 1296.1ab 1835.7ab 1773.3ab 2647.3a 424.7b 1058.2ab 6.7 6.9 851.7 857.5 78.2b 768.6 122.0ab 151.9ab 215.5a 974.2 1389.9 1843.1 45.9ab 111.4b 596.7 1475.3 7.0 860.7 188.0b 253.7b 356.9ab 328.2b 507.6ab 1000.0a 7.2 8.3 8.6 867.0 903.9 917.3 338.9ab 4.8 97.9b 315.1ab 4.2 109.0ab 402.4a 3.4 175.2a 328.3a 3.2 205.6a 175.7b 5.1 55.2b 249.8b 4.5 89.8b 9.2 938.2 4.4b 13.9b 55.0a 53.2a 11.7b 14.1b 10.7 10.8 984.4 989.0 15.1a 31.5 11.8a 39.6 14.9a 13.9 11.2a 9.2 5.0b 24.8 2.6b 11.4 11.3 1004.5 6.6a 5.2a 6.0a 4.8a 1.4b 0.8b 12.1 1032.9 12.6 1049.3 36.2 279.3a 49.4 309.8a 56.1 349.0a 46.0 276.7a 55.8 162.1b 58.2 223.9b 13.3 1072.3 13.4 1075.1 20.1a 41.9a 20.0a 27.8b 20.5a 61.4a 17.9a 35.5b 16.1b 16.8d 16.3b 18.8c 13.8 1086.9 154.7a 144.0a 49.4b 32.3b 40.3b 10.1b 14.1 1095.7 14.3 1100.8 2.1a 4.7b 1.9a 4.9b 1.3a 13.1a 1.4a 11.2ab 0.0b 15.1a 0.0b 15.6a 204.5a 2.9 0.0b 94.9ab 142.4b 3.8 0.0b 68.0b 192.9a 6.2 0.0b 202.6a 25.9d 4.5 2.0a 166.2ab 30.6c 5.1 2.1a 69.7b 0.0b 22.9a 0.0c 0.0b 44.5a 0.0c 14.4 14.8 14.9 15.0 1105.7 1118.4 1124.8 1128.7 15.3 1137.1 15.5 1145.2 15.7 1151.3 16.1 1165.4 16.3 1172.1 3.6ab 2.2 2.5bc 3.7 127.2b 3.7 0.0b 69.7b C6.5 0.9b 21.4ab 3.8ab 0.5b 24.1ab 2.7b 2.5a 14.0b 5.0a 2.1a 9.8b 4.3a 5.8a 3.6 3.5ab 2.8 0.0c 3.4 0.0c 1.7 16.5 1177.6 22.2c 18.8c 39.3ab 27.0bc 39.3ab 46.0a 17.0 1194.6 17.4 1208.3 17.9 1229.9 0.3b 19.6a 72.6 0.2b 20.6a 61.7 0.6b 22.8a 56.7 0.6b 18.9a 73.7 11.4a 13.7b 72.1 8.5a 16.1b 79.4 18.2 1243.5 102.8b 159.9b 202.0ab 177.3ab 214.2a 219.3a 19.7 1301.8 2.8b 3.9b 9.9b 6.5b 13.0ab 14.1a (Continued ) Vol. 75, Nr. 4, 2010—JOURNAL OF FOOD SCIENCE S177 S: Sensory & Food Quality Volatile compound RT4 (min) Maturity stages of pac choi . . . Table 3 --- Continued. Volatile compound 42 2-undecanone 43 Dodecanal 44 6,10-dimethyl-5,9undecadien-2-one (geranylacetone) 45 2-isothiocyanatoethylbenzene 46 Butylated hydroxytoluene 47 Lilial 48 Cedrol Literature aroma Green sour citrus (Hashizume and Samuta 1997; Thi Minh Tu and others 2002) Pungent floral sweet green magnolia-like (Chida and others 2004) RT4 (min) Treatments1,2,3 RI5 O2.5 C2.5 O4.5 C4.5 O6.5 C6.5 19.8 1305.5 22.8 1410.8 0.8a 1.3b 0.8a 1.8b 0.6a 6.4ab 0.6a 10.3a 0.0b 5.2ab 24.0 1457.2 5.6b 10.6ab 26.1ab 16.8ab 34.6ab 41.4a 65.6 64.5 39.7 26.0 36.7 24.4 1476.4 41.0 0.0b 5.3ab 25.5 1518.8 1.2b 6.4b 9.1a 8.4a 7.6a 7.3a 25.8 1534.5 27.7 1615.0 4.0 1.1b 4.2 1.5ab 5.2 2.8ab 3.8 2.2ab 3.8 3.2a 5.5 3.0a 1 O2.5 = Organic at 2.5 wk maturity; C2.5 = Conventional at 2.5 wk maturity; O4.5 = Organic at 4.5 wk maturity; C4.5 = conventional at 4.5 wk maturity; O6.5 = Organic at 6.5 wk maturity; C6.5 = Conventional at 6.5 wk maturity. 2 Concentration of volatile shown in pg per g of pac choi. 3 Different letters (a, b, c) show differences between treatments. 4 Retention time in minutes. 5 Retention index (Kovats) calculated from DB5 column. S: Sensory & Food Quality (Z)-3-hexenal (9) was reported as providing aromas reminiscent of “green,” “green leaves,” and “grassy” in virgin olive oil (Morales and others 1995; Aparicio and others 1997). This compound was also described as providing “strong green” characteristics in green olives (Iraqi and others 2005). Similarly, (E)-2-hexenal (11) was described as being present in blackberries providing “fruit,” “orange,” and “green” aroma characteristics (Klesk and Qian 2003). The same compound was reported as present in virgin olive oil providing “bitter” characteristics (Aparicio and others 1997). (Z)-3-hexen-1-ol (12) was reported as providing “green” aromas in a study focusing on describing sensory characteristics of musty compounds in foods (Chambers and others 1998). Iraqi and others (2005) reported that (Z)-3-hexen-1-ol (12) provided “vanilla” and “green” characteristics in green olives. (E,E)-2,4-hexadienal (15) was identified in fish oil enriched milk and reported to provide “green” and “vegetable” aromas (Venkateshwarlu and others 2004). The same compound had been previously found in mayonnaise and was described as “green” and “burnt” (Hartvigsen and others 2000). Other compounds found in pac choi at lower concentrations that have been previously reported as having “green” characteristics are 1-penten-3-ol (4), octanal (19), (E)-2-octenal (20), (Z)-6-nonenal (34), 2-undecanone (42), acetic acid octyl ester (octyl acetate) (31), and cedrol (48) (Aparicio and others 1997; Buettner and Schieberle 2001; Thi Minh Tu and others 2002; Klesk and Qian 2003; Chida and others 2004; Beaulieu 2005). In a study that focused on the chemicals associated with green odors and flavors in foods, several aldehydes, alcohols, ketones, azoles, and ester derivatives were reported as responsible for the green aroma in foods (Hongsoongnern and Chambers 2008a). The same study reported that the “green” characteristics in foods can be of various types such as unripe, peapod, grassy/leafy, viney, fruity, or may appear as a combination of these. Benzeneacetaldehyde (21) was identified in extruded Amilo rye and described as “flower,” “honey,” and “bitter almond” (Heiniö and others 2003). Interestingly, benzeneacetaldehyde was also described as “green” at a lower intensity in the same study. In addition, 4,5-dimethylthiazole (30) was identified in fried beef steaks and was described as having “smoky,” “roasty,” “fragrant,” and “nutty” aroma characteristics (Specht and Baltes 1994). Other compounds found in pac choi at low concentrations were 2-butanone (1) described as “fragrant” and “pleasant” (Morales and others 1995); 1-octen-3-ol (17) and 1-butene-4-isothiocianato (18) described as “mushroom” and “sulfur,” respectively (Engel and S178 JOURNAL OF FOOD SCIENCE—Vol. 75, Nr. 4, 2010 others 2002); heptanal (14) and 2-nonanone (25) described as “floral” as well as 1-nonanol (29) and 2-decanone (37), which were previously described as “fatty” (Yang and others 2008); camphor (33) described as “piney” and “spicy” (Mehinagic and others 2006); and dodecanal (43), which was previously described as having “citrus” and “skin-like” characteristics (Hashizume and Samuta 1997). Correlating sensory and chemical data Partial least squares (PLS2) regression was used to correlate sensory and chemical data (Figure 1). The analysis showed that 85% of the chemical data explains 86% of the sensory data. The samples that were harvested late (at 6.5 wk) are more correlated with attributes such as overall green, green-grassy/leafy, and salty. The volatile compounds related to these attributes are (Z)-3-hexen-1-ol (12), octyl acetate (31), 1-nonanol (29), and 2decanone (37). These volatiles were present at higher concentrations in the pac choi harvested at 6.5 wk. In fact, 1-nonanol (29) was only present in these samples and not in the samples harvested earlier. These chemicals have been associated with “bitter,” “green,” “fruity,” and “fatty” aromatics (Aparicio and others 1997; Thi Minh Tu and others 2002; Iraqi and others 2005; Yang and others 2008). Other compounds which are also closely associated with samples harvested at 6.5 wk are 1-penten-3-ol (4), 3,7-dimethyl-1,6-octadien-3-ol (linalool) (26), camphor (33), menthol (36), isobornyl acetate (41), 6,10-dimethyl-5,9-undecadien-2one (geranylacetone) (44), and cedrol (48). These chemicals have also been associated with “green,” “floral,” “woody,” “citrus,” and “piney” aromatics (Chida and others 2004; Mehinagic and others 2006). In our study, these compounds were closely related to attributes such as bitter, toothetch, soapy, cabbage, sulfur, and woody. Fiber awareness and crispness are textural attributes also related to these samples and these volatiles. This may indicate colinear attributes that change similarly but have different etiologies. The spinach flavor attribute was related to with dodecanal (43), an aldehyde with citrus aromatics that has also been found in cilantro and carrots in the past (Buttery and others 1968; Hashizume and Samuta 1997; Fan and Sokorai 2002). Similarly, butylated hydroxytoluene (BHT) (46) was closely related to the spinach flavor of pac choi. Parsley flavor was related to butyl isothiocyanate (1-isothiocyanato-butane) (16), a derivative from glucosinolates, which are frequently found in vegetables from the Brassica family and more specifically cabbage (Ciska and Pathak compound which is also closely related to samples at their early stage of development is benzyl nitrile (benzene acetonitrile) (32), which is another chemical formed from the degradation of glucosinolates. This compound was previously identified and quantified in turnip greens at different stages of development (Jones and others 2007). Those authors found that benzene acetonitrile actually increased as the plant got older. However, the concentrations were generally small. The pac choi samples harvested at an early stage of growth (2.5 wk) were generally associated with green-unripe, piney, and musty/earthy flavors as well as moistness. The concentrations of many volatiles varied among maturity levels of pac choi. In addition, differences also are noted between organically and conventionally grown pac choi for a few volatiles. The volatiles that were generally higher for conventionally grown pac choi were (Z)-3-hexenal (9), 2-hexyn-1-ol (10), and (E)-2-hexenal (11). These compounds are responsible for the “green” and “bitter” aroma in foods (Aparicio and others 1997). This is in agreement with the sensory analysis of pac choi which showed that conventional pac choi had significantly higher intensities of overall green, bitter, and soapy attributes compared to organic pac choi at the earliest stage of development (2.5 wk). However, the intensities of overall green, bitter, and soapy are similar between organic and conventional pac choi at both 4.5 and 6.5 wk maturity levels. It may be that the introduction of other flavor volatiles such as 1-penten3-ol (4), linalool (26), and geranylacetone (44) is balancing the perception of overall green and bitter at later stages of growth. These compounds also have been associated with “green” and “floral” aromas in the past (Chida and others 2004). Butanoic acid methyl ester (5) and 4,5-dimethylthiazole (30) were generally higher in organic pac choi. However, these differences did not translate in sensory flavor differences between organic and conventional pac choi. Other compounds that were higher for organic pac choi were 2-methyl-3-pentanone (6), 1-octanol (23), (E)-2-nonen-1-ol (27), camphor (33), and (Z)-6-nonenal (34). However, these chemicals were present at low concentrations. In summary, the differences in volatile compounds among stages of growth are more substantial compared to the differences between organic and conventional production systems. In many cases, these differences in chemical composition do translate into the flavor characteristics observed in pac choi. 2004). In another study, butyl isothiocyanate (16) was also identified in cooked cauliflower and was described as having “sulfur,” “green,” and “pungent” aroma characteristics (Engel and others 2002). Other volatile compounds closely related to the parsley flavor were 2-hexyn-1-ol (10) and (E,E)-2,4-hexadienal (15). It has been suggested that several aldehydes, alcohols, ketones, or ester derivatives with 6 carbon atoms (C6 ) in their molecules are responsible for the “green” aroma in foods (Hongsoongnern and Chambers 2008a). (E,E)-2,4-hexadienal (15) has been previously described as having “ripe fruit,” “green,” and “vegetable” aroma characteristics (Aparicio and others 1997; Venkateshwarlu and others 2004). The pac choi samples harvested at 4.5 wk also were correlated to the parsley and spinach flavors. This means that samples harvested at 4.5 and 6.5 wk were usually rated at a higher intensity for these flavors compared to the samples harvested at 2.5 wk of maturity. Another group of volatiles were related with the lettuce, umami, and overall sweet attributes. This suggests that these volatiles may have “green” characteristics that are less intense compared to other chemicals more closely associated with parsley, green-grassy/leafy, and overall green attributes. The chemicals associated with lettuce, umami, and sweet flavors are (Z)-3-hexenal (9), octanal (19), benzeneacetaldehyde (21), 4-methyl-1-undecene (22), 1-octanol (23), 2-nonanone (25), (E)-2-nonen-1-ol (27), (Z)-6-nonenal (34), (E)-2decen-1-ol (38), and 2-undecanone (42). This is in agreement with past studies in which many of these compounds have been described as having “sweet,” “floral,” “citrus,” “fruity,” and “green” characteristics. For example, (Z)-3-hexenal (9), octanal (19), and 2undecanone (42) were described as “green” (Aparicio and others 1997; Buettner and Schieberle 2001; Klesk and Qian 2003). At the same time, octanal (19) has also been described as having “sweet” and “citrusy” aroma characteristics (Thi Minh Tu and others 2002). Benzeneacetaldehyde (21) has been reported as having “flower,” “honey,” “sweet” and “green” aroma characteristics (Heiniö and others 2003). 2-Nonanone (25) was described as “fruity” and “floral” (Yang and others 2008) and (Z)-6-nonenal (34) was reported as “citrus,” “green,” “cucumber,” and “melon-like” (Beaulieu 2005). These compounds are more correlated to pac choi samples harvested at both 2.5 and 4.5 wk maturity. This means that their concentration is higher at early stages and decrease as the plants get older. Another 16 Parsley ID Spinach ID 15 10 Lettuce ID Umami 27 PC2 19 23 9 34 21 38 Sweet, Overall 43 O4.5 C4.5 46 Bitter Toothetch 26 33 Fiber Awareness 48 Crispness 36 Cabbage ID Soapy 41 Sulfur 44 Woody 12 31 4 30 22 42 25 Figure 1 --- Partial least squares regression (PLS) correlating sensory and instrumental data. O2.5 = organic at 2.5 wk maturity; C2.5 = conventional at 2.5 wk maturity; O4.5 = organic at 4.5 wk maturity; C4.5 = conventional at 4.5 wk maturity; O6.5 = organic at 6.5 wk maturity; C6.5 = conventional at 6.5 wk maturity. C6.5 32 Green, Grassy / Leafy O6.5 C2.5 O2.5 Green, Unripe 29 37 Salty Green, Overall Piney Musty / Earthy Moistness Green, Peapod PC1 Vol. 75, Nr. 4, 2010—JOURNAL OF FOOD SCIENCE S179 S: Sensory & Food Quality Maturity stages of pac choi . . . Maturity stages of pac choi . . . Conclusions M any more differences in sensory characteristics and chemical profile are observed among stages of growth of pac choi compared to the production method. Pac choi harvested early (2.5 wk) is described as green-unripe, piney, musty/earthy, and moist. As the plant grows, other flavors such as lettuce, umami, and overall sweet develop. These flavors are correlated with volatiles that have been associated with “sweet,” “floral,” “citrus,” “fruity,” and “green” aromas in the past. These volatiles are (Z)-3-hexenal (9), octanal (19), benzeneacetaldehyde (21), 4-methyl-1-undecene (22), 1-octanol (23), 2-nonanone (25), (E)-2-nonen-1-ol (27), (Z)6-nonenal (34), (E)-2-decen-1-ol (38), and 2-undecanone (42). Finally, when the plant reaches a mature stage at 6.5 wk, it is perceived as having higher intensities of green, bitter, cabbage, sulfur, and woody flavors. These flavors may be associated with the presence of volatiles such as (Z)-3-hexen-1-ol (12), octyl acetate (31), 1-nonanol (29), 2-decanone (37), 1-penten-3-ol (4), linalool (26), camphor (33), menthol (36), isobornyl acetate (41), geranylacetone (44), and cedrol (48), which have been associated with “strong green,” “bitter,” “fruity,” and “fatty” odors in the past. Finally, conventional pac choi was higher in green overall, bitter, and soapy flavors compared to organic pac choi when harvested at 2.5 wk only. 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