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Analysis of glycative products in sauces and sauce-treated foods
Food Chemistry 113 (2009) 262–266
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Analytical Methods
Analysis of glycative products in sauces and sauce-treated foods Pei-chun Chao a,b, Cheng-chin Hsu b, Mei-chin Yin c,* a b c
Department of Nutrition, Chung Shan Medical University Hospital, Taiwan, ROC Department of Nutritional Science, Chung Shan Medical University, Taiwan, ROC Department of Nutrition, China Medical University, 91, Hsueh-shih Road, Taichung City, Taiwan, ROC
a r t i c l e
i n f o
Article history: Received 13 February 2008 Received in revised form 27 June 2008 Accepted 30 June 2008
Keywords: Pentosidine Carboxymethyllysine Soybean sauce Barbecue sauce Maillard reaction
a b s t r a c t Content of Maillard reaction products (MRPs) such as pentosidine, carboxymethyllysine and furosine in soybean sauce, sour-sweet sauce, tomato sauce, barbecue sauce, and sauce-treated chicken, pork, beef, salmon and cod was analysed. In test sauces, MRP content was in the range of 10–692 lg/100 mL sample. MRP content in raw, boiled, fried and baked foods was in the range of 10–76 lg/100 g sample. Boiling, frying and baking caused significantly higher MRP levels in test foods (P < 0.05). MRP levels in soybean sauce, sour-sweet sauce and barbecue sauce-treated foods were in the range of 1094–2424, 1494– 3146 and 1400–2926 lg/100 g sample. The interactions of sauce, heating and frying oil markedly enhanced the formation of MRPs in sauce-treated foods. Because MRPs are glycative products; thus, patients with glycation associated diseases may consider limiting the dietary use of these sauces. Ó 2008 Elsevier Ltd. All rights reserved.
1. Introduction Maillard reaction (MR) occurs between the free amino group of amino acids and the carbonyl group of reducing sugars such as glucose and fructose, and leads to the formation of reversible Amadori products, which spontaneously undergo further nonenzymatic rearrangement to form irreversible crosslinks called Maillard reaction products (MRPs) or advanced glycation endproducts (AGE) (Grandhee & Monnier, 1991; Yaylayan & Huyghues-Despointes, 1994). The common Amadori products or MRPs include pentosidine, carboxymethyllysine (CML) and furosine (Erbersdobler & Somoza, 2007; Goldberg et al., 2004). The presence of these MRPs in food may benefit food preservation because these MRPs could provide antioxidative protection (Bedinghaus & Ockerman, 1995). However, the impact of dietary MRPs upon healthy risk have been considered because dietary MRP content contributes to excess serum AGE levels, and the elevated AGE level in circulation favours the development of glycation and inflammation associated diseases such as renal failure, diabetes and Alzheimer’s disease (Koschinsky et al., 1997; Riviere, Birlouez-Aragon, & Vellas, 1998; Vlassara et al., 2002). Therefore, in order to avoid the adverse effects of dietary MRPs upon physiological variation of AGE levels, and the progression or deterioration of glycative injury, it is important to characterise and quantify MRPs in the diet. Soybean sauce, sour-sweet sauce, barbecue sauce and tomato sauce (ketchup) are less expensive and easily available sauces in * Corresponding author. Tel.: +886 4 22053366x7510; fax: +886 4 22062891. E-mail address: [email protected] (M.-c. Yin). 0308-8146/$ - see front matter Ó 2008 Elsevier Ltd. All rights reserved. doi:10.1016/j.foodchem.2008.06.076
Taiwan and Asia. Soybean sauce is a fermentative product of soybean, rich in protein. Sour-sweet sauce and barbecue sauce have sweet taste because sugar is an ingredient in these sauces. Therefore, these sauces are prone to undergo MR with foods rich in protein during food preparation and storage. Although these sauces are used for seasoning, they are used frequently and in a large amount in some food preparations in Asia. Thus, the influence of these sauces upon consumer’s healthy risk could not be ignored. So far, less information is available regarding the MRP content in these sauces, and sauce-treated foods. This study analysed the content of MRPs such as pentosidine, CML and furosine in commercially available soybean sauce, soursweet sauce, tomato sauce and barbecue sauce as well as foods rich in protein such as chicken, pork, beef, salmon and cod. The influence of boiling, frying or baking treatment upon MRP formation in foods and sauce-treated foods was also determined. The results of this study could provide novel information for people with glycation associated diseases to make appropriate dietary choices. 2. Materials and methods 2.1. Sauce and food sample Soybean sauce, sour-sweet sauce, tomato sauce (ketchup), barbecue sauce, chicken, pork, beef, salmon, and cod were purchased from local supermarkets in Taichung City (Taiwan). For commercial sauces, four brands of each sauce were examined, in which eight samples for each brand were used. Skin and visible fat of chicken, pork, beef, salmon, and cod were removed.
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2.2. Lipid oxidation measurement Food freshness could be evaluated by the level of lipid oxidation (Pantazi, Papavergou, Pournis, Kontominas, & Savvaidis, 2008). Thus, in order to ensure the freshness of chicken, pork, beef, salmon, and cod, lipid oxidation level of these food samples was determined by a thiobarbituric acid (TBA)-assay described by Schmedes and Hølmer (1989), in which 10 g sample were chopped and mixed with 25 mL 20% trichloroacetic acid solution (200 g/L of trichloroacetic acid in 135 mL/L phosphoric acid solution) and homogenised in a Waring Blender for 30 s. After filtration, a 2mL filtrate was combined with 2 mL, 0.02 M aqueous TBA in a test tube. The tubes were incubated at room temperature in the dark for 20 h, and the absorbance at 532 nm was measured by using UV–VIS spectrophotometer (model UV-1210, Shimadzu, Tokyo, Japan). Thiobarbituric acid reactive substances (TBARS) content were expressed as malondialdehyde equivalents (mg/kg food sample). In this study, malondialdehyde equivalents of used samples were in the range of 0.06–0.091 mg/kg food sample.
CML by reverse phase HPLC. Pentosidine was analysed by an HPLC method described by Miyata et al. (1996), in which a Supelco reverse phase C18 column (id, 4.6 mm; od, 25 cm; Supelco, Bellefonte, PA, USA) were equipped. The effluent was monitored by a fluorescence detector (RF-10A Shimadzu, Kyoto, Japan), and excitation and emission wavelength were set at 335 and 385 nm, respectively. The content of CML and pentosidine in used soybean oil, water and sauce for soaking, boiling or frying were analysed. CML and pentosidine levels in these samples were lower than 10 lg/100 mL. 2.6. Determination of furosine
The content of reducing sugar (glucose and fructose) in sauces and food samples was determined according to the method described by Ohara-Takada et al. (2005). Briefly, 10 mL sauce or 10 g food sample were mixed with 50 mL 80% ethanol. After 1 h heating at 80 °C, sample was cooled down to room temperature, and followed by centrifugation (2500g for 15 min at 25 °C). Supernatant was collected, dried under vacuum, re-dissolved in distilled water and passed through a 0.2-lm membrane filter. The concentration of glucose and fructose in filtrate was analysed by an HPLC (Shimadzu LC-10, Kyoto, Japan) quipped with an evaporative light scattering detector (Sedex, Paris, France) and a TSK gel Amide-80 column (id, 4.6 mm; od, 25 cm; Anachem, Luton, UK).
Furosine was determined according to the method described by Delgado, Corzo, Santa-Maria, Jimeno, and Olano (1992). Food or sauce-treated food, 50 g, was homogenised in 50 mL sterile distilled water. After filtrating through a Whatman No. 1 filter paper, the filtrate was collected. Then, 50 mL sauce or food filtrate were hydrolysed, using 3 mL of 7.95 M HCl at 110 °C for 23 h in screwcap tubes. Hydrolysis tubes were sealed under nitrogen. Hydrolysates were aerated and cooled down, and followed by centrifugation at 14,000g for 10 min. The supernatant was applied to a SepPak C18 cartridge (Millipore, Milfold, MA, USA) pre-wetted with 5 mL methanol and 10 mL deionized water, and followed by eluting with 3 mL 3 M HCl. The dried sample was re-dissolved in 1 mL mixture containing water, acetonitrile and formic acid at the ratio of 95:5:0.2 (v/v/v). Degassed mobile phase was prepared with 5 mM sodium heptane sulphonate including 20% acetonitrile and 0.2% formic acid. Furosine was quantified by the external standard method. Calibration curve was built from a stock solution in the range of 1–2000 lg/100 mL. The content of furosine in used soybean oil, water and sauce for soaking, boiling or frying were analysed. Furosine level in these samples was lower than 14 lg/ 100 mL.
2.4. Food preparation
2.7. Statistical analysis
In Asia, tomato sauce is not commonly used for cooking; thus, this sauce was not used to treat any food in this study. Each sauce used for food treatment was mixed from four commercial brands at equal amounts. Foods and sauce-treated foods were prepared as follows. For boiling preparation, 200 g food were cut into 4 strips and mixed with 400 mL sauce (or water), and boiled at 100 °C for 15 min. For deep frying preparation, 200 g food were cut into 4 strips and soaked with 400 mL sauce (or water) for 2 h at 25 °C. Then, sauce (or water) residue on food surface was removed by paper towel, and food sample was treated by deep frying in 100 mL soybean oil at 180 °C for 15 min. For baking preparation, 200 g food were cut into 4 strips and soaked with 400 mL sauce (or water) for 2 h at 25 °C. Then, sauce (or water) residue on food surface was removed by paper towel, and food sample was treated by baking at 230 °C for 15 min. After prepared, food was put on a Whatman No. 1 filter paper for 30 min for cooling and removing the residue of sauce or frying oil.
The effect of each treatment was analysed from eight different preparations (n = 8). Data were reported as means ± standard deviation (SD), and subjected to analysis of variance (ANOVA). Differences among means were determined by the least significance difference test with significance defined at P < 0.05.
2.3. Measurement of reducing sugars
2.5. Determination of carboxymethyllysine (CML) and pentosidine The method described by Inagi et al. (2006) was used to determine CML level. Sauce sample, 50 mL, or food and sauce-treated food sample, 50 g, was mixed with excess of NaBH4 in 0.2 mol/L borate buffer (pH 9.1) for reduction. Proteins were precipitated by 20% trichloroacetic acid, and centrifugation at 2000g for 10 min. The pellet was washed with 1 mL 10% trichloroacetic acid. After drying, the pellet was acid hydrolysed in 500 lL 6 M HCl for 16 h at 110 °C in screw-cap tubes purged with nitrogen. Hydrolysates were dried, and rehydrated in water, and used for measuring
3. Results Content of protein, lysine, glucose and fructose in soybean sauce, sour-sweet sauce, tomato sauce, barbecue sauce and food
Table 1 Content of protein (g), lysine (mg), glucose (mg) and fructose (mg) in 100 mL or 100 g soybean sauce, sour-sweet sauce, tomato sauce, barbecue sauce, chicken, pork, beef, salmon and cod Protein
Lysine
Glucose
Fructose
Soybean sauce Sour-sweet sauce Tomato sauce Barbecue sauce
5.2 1.6 1.2 3.6
204 –e – 178
16 ± 4a 37 ± 7c 10 ± 5a 23 ± 8b
7 ± 2a 58 ± 8c 41 ± 6b 76 ± 11d
Chicken Pork Beef Salmon Cod
13.4 12.4 9.8 13.0 8.8
1452 1868 1728 1626 1540
– – – – –
– – – – –
Data are mean ± SD, n = 8. Protein and lysine data are from Taiwan Nutrient Database. a–d Means in a column without a common letter differ, P < 0.05. e Means too low to be detected.
4. Discussion Our present study found 4 sauces contained furosine, CML and pentosidine. Furosine is a degradation product of Amadori compounds; CML and pentosidine are irreversible crosslinks formed from Amadori products during advanced stages of Maillard reactions. Therefore, the presence of these three MRPs strongly implied that Maillard reaction has occurred, and might be still undergoing in these sauces. Furthermore, we found the content of pentosidine, CML and furosine in each type of sauce was from brand to brand. It is known that the MRPs content in foods is related to their composition, the method for processing and condition for storage (Chuyen, 2006; Goldberg et al., 2004). Because these sauces contained various levels of protein, lysine and reducing sugars (glucose and
Table 2 Content (lg/100 mL sample) of pentosidine, CML and furosine in soybean sauce (SB), sour-sweet sauce (SS), tomato sauce (TS) and barbecue sauce (BB) Sauce
MRP
Range
Mean (n = 8)
SB
Pentosidine CML Furosine
10–82 172–392 28–102
42 ± 18 224 ± 43 58 ± 26
SS
Pentosidine CML Furosine
24–104 124–532 24–110
66 ± 25 306 ± 63 74 ± 19
TS
Pentosidine CML Furosine
20–76 176–692 42–120
52 ± 16 416 ± 89 60 ± 21
BB
Pentosidine CML Furosine
78–158 252–482 26–110
104 ± 46 352 ± 70 76 ± 17
90
d
70
d
d
d
60
raw
c c
c c
50
c
b
boiled
b
b
40
fried
b
30 20
e
e
80
a
baked
a
a
a
a
10 0 chicken
pork
90
CML (ug/100 g sample)
samples is shown in Table 1. Food samples had more protein and lysine than sauces; however, sauces had more glucose and fructose than food samples. The content of glucose in sauces followed the order: sour-sweet sauce > barbecue sauce > soybean sauce = tomato sauce (P < 0.05). Fructose content in sauces followed the order: barbecue sauce > sour-sweet sauce > tomato sauce > soybean sauce (P < 0.05). The content of pentosidine, CML and furosine in soybean sauce, sour-sweet sauce, tomato sauce, and barbecue sauce is presented in Table 2. The contents of these three MRPs were different from brand to brand, and in the range of 10– 692 lg/100 mL sauce. The content of pentosidine, CML and furosine in raw, boiled, fried or baked chicken, pork, beef, salmon and cod is shown in Fig. 1. In test foods, pentosidine, CML and furosine contents were in the range of 10–72, 14–76, and 22–68 lg/ 100 g food. The process of boiling, frying and baking significantly increased levels of pentosidine, CML and furosine in test foods (P < 0.05). In salmon and cod, frying process caused significantly higher levels of pentosidine and CML than boiling or baking process (P < 0.05). The content of pentosidine, CML, furosine and sum of these three glycative products in boiled, fried and baked chicken, pork, beef, salmon and cod treated by soybean sauce, sour-sweet sauce or barbecue sauce is shown in Tables 3–5, respectively. The sum of pentosidine, CML and furosine levels in soybean sauce-treated foods were in the range of 1094–2424 lg/100 g sample. Soursweet sauce-treated foods had total level of pentosidine, CML and furosine in the range of 1494–3146 lg/100 g sample. Barbecue sauce-treated foods had total level of pentosidine, CML and furosine in the range of 1400–2926 lg/100 g sample. For each test food sample, frying and baking treatments caused significantly greater formation of pentosidine, CML and furosine than boiling treatment (P < 0.05).
pentosidine (ug/100 g sample)
P.-c. Chao et al. / Food Chemistry 113 (2009) 262–266
beef
salmon
cod
d
d
d
d
80 70
c
c
c
c c
c
60 50
b
b
b
b
b
boiled fried
40
baked
a
30 20
raw
a
a a
a
10 0 chicken
pork
80
furosine (ug/100 g sample)
264
beef
salmon
cod
d
d d
70 c 60
c
c
c
c
c
c c c
c
b
50
b
40 a
fried
a
30
raw boiled
a a
a
baked
20 10 0 chicken
pork
beef
salmon
cod
Fig. 1. Content (lg/100 g sample) of pentosidine, CML and furosine in raw, boiled, fried or baked chicken, pork, beef, salmon and cod. Data are mean ± SD (n = 8). (a–e) Means among bars without a common letter differ, P < 0.05.
Table 3 Content (lg/100 g sample) of pentosidine, CML and furosine in boiled, fried and baked chicken, pork, beef, salmon and cod treated by soybean sauce Sample
Method
Pentosidine
CML
Furosine
Sum
Chicken
Boiled Fried Baked
192 ± 24 206 ± 29 254 ± 23
638 ± 45 878 ± 48 694 ± 41
688 ± 49 752 ± 40 900 ± 47
1518 ± 52c 1836 ± 68d 1848 ± 74d
Pork
Boiled Fried Baked
174 ± 20 260 ± 25 324 ± 30
572 ± 48 904 ± 44 922 ± 60
840 ± 51 692 ± 38 962 ± 63
1526 ± 66c 1856 ± 60d 2208 ± 87e
Beef
Boiled Fried Baked
182 ± 18 291 ± 31 340 ± 27
526 ± 49 854 ± 61 764 ± 42
820 ± 40 654 ± 25 776 ± 43
1538 ± 69c 1798 ± 80d 1880 ± 93d
Salmon
Boiled Fried Baked
148 ± 20 238 ± 29 256 ± 31
486 ± 47 980 ± 50 1078 ± 65
460 ± 32 1206 ± 72 954 ± 59
1094 ± 55a 2424 ± 84f 228 ± 83e
Cod
Boiled Fried Baked
160 ± 19 218 ± 33 270 ± 27
550 ± 58 846 ± 76 952 ± 64
544 ± 44 858 ± 62 688 ± 50
1254 ± 71b 1922 ± 89d 1910 ± 73d
Data are mean ± SD, n = 8. a–f Means in a column without a common letter differ, P < 0.05.
fructose); thus, MRPs could be formed naturally in these sauces during storage. In addition, the results of our present study revealed that fried salmon and cod had higher pentosidine and CML levels than baked samples, although the baking temperature (230 °C) was higher than frying temperature (180 °C). It was notified that heat directly acted on food during the baking process; however, during frying process, heat acted on frying oil first and food obtained heat from the hot frying oil. Thus, MRPs presented in fried food may comprise
P.-c. Chao et al. / Food Chemistry 113 (2009) 262–266 Table 4 Content (lg/100 g sample) of pentosidine, CML and furosine in boiled, fried and baked chicken, pork, beef, salmon and cod treated by sour-sweet sauce Sample
Method
Pentosidine
CML
Furosine
Sum
Chicken
Boiled Fried Baked
152 ± 20 190 ± 31 334 ± 33
906 ± 57 1350 ± 76 1486 ± 81
512 ± 38 462 ± 34 680 ± 47
1570 ± 62a 2002 ± 75c 2500 ± 90c
Pork
Boiled Fried Baked
164 ± 23 226 ± 36 310 ± 29
808 ± 66 1242 ± 79 1426 ± 91
522 ± 45 668 ± 56 604 ± 53
1494 ± 73a 2136 ± 83c 2340 ± 104d
Beef
Boiled Fried Baked
186 ± 27 254 ± 42 362 ± 40
870 ± 71 1364 ± 83 1208 ± 68
527 ± 32 790 ± 60 684 ± 55
1578 ± 77a 2408 ± 92e 2254 ± 73d
Salmon
Boiled Fried Baked
170 ± 25 302 ± 29 276 ± 34
1084 ± 59 1670 ± 86 2180 ± 101
528 ± 34 924 ± 56 690 ± 49
1782 ± 70b 2896 ± 92g 3146 ± 120h
Cod
Boiled Fried Baked
154 ± 19 188 ± 32 258 ± 39
974 ± 78 1522 ± 89 1924 ± 91
612 ± 59 1064 ± 88 766 ± 76
1740 ± 92b 2774 ± 95f 2948 ± 112g
Data are mean ± SD, n = 8. a–h Means in a column without a common letter differ, P < 0.05.
Table 5 Content (lg/100 g sample) of pentosidine, CML and furosine in boiled, fried and baked chicken, pork, beef, salmon and cod treated by barbecue sauce Sample
Method
Pentosidine
CML
Furosine
Sum
Chicken
Boiled Fried Baked
452 ± 46 490 ± 56 634 ± 43
542 ± 47 844 ± 63 1152 ± 87
406 ± 45 568 ± 60 778 ± 66
1400 ± 73a 1902 ± 82c 2564 ± 115f
Pork
Boiled Fried Baked
460 ± 38 534 ± 57 602 ± 64
620 ± 36 882 ± 65 1226 ± 71
680 ± 60 736 ± 65 646 ± 54
1760 ± 74b 2152 ± 90d 2474 ± 97f
Beef
Boiled Fried Baked
486 ± 55 562 ± 58 672 ± 67
588 ± 69 790 ± 78 1168 ± 95
456 ± 38 534 ± 50 650 ± 63
1530 ± 74a 1886 ± 82c 2490 ± 103f
Salmon
Boiled Fried Baked
440 ± 46 614 ± 70 546 ± 58
632 ± 62 1030 ± 92 1522 ± 106
618 ± 65 1144 ± 85 858 ± 60
1690 ± 76b 2788 ± 94g 2926 ± 88h
Cod
Boiled Fried Baked
434 ± 35 508 ± 49 528 ± 61
546 ± 57 828 ± 86 1506 ± 92
522 ± 43 980 ± 76 704 ± 54
1502 ± 70a 2316 ± 91e 2738 ± 108g
Data are mean ± SD, n = 8. a–h Means in a column without a common letter differ, P < 0.05.
of MRPs from both heat-treated frying oil and frying oil-treated food. We also analysed the content of CML and pentosidine in used soybean oil and found that CML and pentosidine levels in these samples were lower than 10 lg/100 mL. Thus, the MRPs presented in fried foods in our present study were mainly from the interaction of hot frying oil and food. Soybean oil was used in this study for frying treatment. This oil and two test fish are rich in unsaturated fatty acids. It has been reported that MRPs could be formed from Schiff base and Amadori adducts by autoxidation of fatty acids and amino acids (Bucala, Makita, Koschinsky, Cerami, & Vlassara, 1993; Fu et al., 1996). Zamora and Hidalgo (2005) also indicated that lipid oxidation and Maillard reaction were interrelated, and lipid oxidation products could affect Maillard pathway. Thus, it is possible that heat caused oxidation of unsaturated fatty acids from soybean oil and fish, and the oxidative products participated the rearrangement of Amadori adducts to form irreversible crosslinks like pentosidine or CML. Furthermore, we also notified that fried or baked sauce-treated salmon had markedly higher sum of CML, pentosidine and furosine than fried or baked sauce-treated cod. Salmon has 54.9% monounsaturated fatty acids (MUFA) and 24.3% polyunsaturated fatty acids (PUFA); how-
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ever, cod has 73.1% MUFA and 6.1% PUFA (Department of Health, 1998). Obviously, the role of PUFA on MRP formation can not be ignored. These findings suggest that frying or baking treatment should be limited for foods rich in unsaturated fatty acids in order to reduce MRPs formation in foods. Furosine is a direct marker of lysine reaction product and also a relevance for nutritionally unavailable for lysine from food source (Bujard & Finot, 1978; Del Castillo, Corzo, & Olano, 1999). Lysine content of test foods in our present study was in the range of 1452–1868 mg/100 g (Department of Health, 1998). Our present study found the furosine levels of sauce and foods treated by boiling, frying or baking were in the range of 24–120 and 24–68 lg/ 100 mL or 100 g, respectively. However, furosine level in saucetreated foods was in the range of 406 (boiled barbecue sauce-treated chicken)–1206 (fried soybean sauce-treated salmon) lg/100 g. Obviously, about 200–1000 lg/100 g newly produced furosine resulted from MR during food preparation and processing, in which lysine from foods was involved. After calculation, we found that at least 100–540 lg lysine/100 g sauce-treated foods (equal to 0.8–4.5 mg lysine/100 g protein) was nutritional unavailable. Pentosidine could be formed from arginine and lysine (Grandhee & Monnier, 1991). Thus, the formation of pentosidine in sauce-treated foods as we observed also indicated not only thermal damage in food protein but also loss of nutritional available arginine or lysine. Fu et al. (1996) reported that CML could be produced by many pathways such as degradation of the Amadori compounds or direct reaction of lysine with some sugars or lipid oxidation derived dicarbonyl compounds. Erbersdobler and Somoza (2007) indicated that in more severely heat-treated food items, in which furosine levels have already decreased, CML can provide additional information on the protein damage. In our present study, the CML level in baked sour-sweet sauce-treated salmon and cod was very high (2180 and 1924 lg/100 g sample). This finding revealed that advanced MR had occurred in these food samples and also implied that these foods had been severely heat-treated, in which baking treatment provided heat, sour-sweet sauce provided reducing sugars, salmon (or cod) provided lysine and/or unsaturated fatty acids. Thus, CML formation in these food samples involved many factors and complicated chemical reactions. Therefore, in order to maintain protein quality of food and decrease the production of MRPs including CML, pentosidine and furosine, it is better to reduce using baking process and/or sauces for food preparation. On the other hand, we found sour-sweet sauce contained more reducing sugars than soybean sauce, and sour-sweet sauce-treated foods had more MRPs than soybean sauce-treated foods. This finding agreed that reducing sugar is an important factor contributed to MRPs formation, and also partially explains the impact of sauce upon MRP formation in sauce-treated foods. In addition, pentosidine (or CML) level in each sauce-treated food was higher than the sum of pentosidine (or CML) from corresponding sauce, and pentosidine (or CML) from corresponding food. For instance, the CML level of soybean sauce and boiled chicken were 224 lg/ 100 mL and 30 lg/100 g, respectively. However, the CML level in boiled soybean sauce-treated chicken was 638 lg/100 g. Also, the pentosidine level of barbecue sauce and fried salmon was 104 lg/100 mL and 70 lg/100 g only; but, the pentosidine level of fried barbecue sauce-treated salmon was 614 lg/100 g. These results revealed that sauce and heat caused a synergistic effect on MRPs formation in these sauce-treated foods. It is highly possible that heat caused more amino acids release from the tested foods, which allowed reducing sugars from sauces to react with the released amino acids and facilitated the formation of MRPs. Tomato sauce, or so-called ketchup, is commonly used for foods popular in western society such as French fried, hamburger and hot dog. Soybean sauce, sour-sweet sauce, and barbecue sauce are popular in Asia for food preparation. Although they are commonly used
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for seasoning only, they are used with high frequency and large quantity in certain food preparations. In Taiwan and Japan, soybean sauce could be directly used as table sauce like ketchup for foods such as tofu and sashimi (raw fish). It has been indicated that MRPs possess antioxidative property and could extend shelf-life of food via delaying lipid oxidation (Bedinghaus & Ockerman, 1995; Wijewickreme & Kitts, 1998); thus, these MRPs might benefit food preservation. However, the dietary MRPs could elevate AGE levels in circulation, which consequently contributes to the progression of glycation associated diseases such as diabetes, Alzheimer’s disease and inflammatory arthritis (Uribarri et al., 2007). Thus, patients with glycation associated diseases should avoid consuming foods or sauces rich in MRPs based on healthy consideration. In conclusion, soybean sauce, sour-sweet sauce, tomato sauce, barbecue sauce contained reducing sugars, and the content of pentosidine, CML and furosine in these sauces was in the range of 10–692 lg/100 mL. MRP content in sauce-treated foods was in the range of 1094–3146 lg/100 g. Besides heat, frying oil played a role to affect the formation of MRPs in test foods. Furthermore, sauce and heat caused a synergistic effect in the formation of MRPs in sauce-treated foods. These data suggest that patients with glycation associated diseases should consider limiting the dietary use of these sauces. References Bedinghaus, A. J., & Ockerman, H. W. (1995). Antioxidative Maillard reaction products from reducing sugars and free amino acids in cooked ground pork patties. Journal of Food Science, 60, 992–995. Bucala, R., Makita, Z., Koschinsky, T., Cerami, A., & Vlassara, H. (1993). Lipid advanced glycosylation: Pathway for lipid oxidation in vivo. Proceedings of the National Academy of Sciences of the United States of America, 90, 6434–6438. Bujard, E., & Finot, P. A. (1978). Measure of available and blocked lysine in industrial milks. Annales de la nutrition et de l’alimentation, 32, 291–305. Chuyen, N. V. (2006). Toxicity of the AGEs generated from the Maillard reaction: On the relationship of food-AGEs and biological-AGEs. Molecular Nutrition and Food Research, 50, 1140–1149. Del Castillo, D. M., Corzo, N., & Olano, A. (1999). Early stages of Maillard reaction in dehydrated orange juice. Journal of Agricultural and Food Chemistry, 47, 4388–4390. Delgado, T., Corzo, N., Santa-Maria, G., Jimeno, M. L., & Olano, A. (1992). Determination of furosine in milk samples by ion pair reversed phase liquid chromatography. Chromatographia, 33, 374–376. Department of Health (1998). Taiwan nutrient databases 1998. Taipei, Taiwan: Department of Health, Executive Yuan. Erbersdobler, H. F., & Somoza, V. (2007). Forty years of furosine – forty years of using Maillard reaction products as indicators of the nutritional quality of foods. Molecular Nutrition and Food Research, 51, 423–430.
Fu, M. X., Requena, J. R., Jenkins, A. J., Lyons, T. J., Baynes, J. W., & Thorpe, S. R. (1996). The advanced glycation endproduct Nepsilon-[carboxymethyl]-lysine, is a product of both lipid peroxidation and glycoxidation reactions. Journal of Biological Chemistry, 271, 9982–9986. Goldberg, T., Cai, W., Melpomeni, P., Dardaine, V., Baliga, B. S., Uribarri, J., et al. (2004). Advanced glycoxidation end products in commonly consumed foods. Journal of American Dietician Association, 104, 1287–1291. Grandhee, S. K., & Monnier, V. M. (1991). Mechanism of formation of the Maillard protein cross-link pentosidine. Journal of Biological Chemistry, 266, 11649–11653. Inagi, R., Yamamoto, Y., Nangaku, M., Usuda, N., Okamato, K., Kurokawa, K., et al. (2006). A severe diabetic nephropathy model with early development of nodule-like lesions induced by megsin overexpression in RAGE/iNOS transgenic mice. Diabetes, 55, 356–366. Koschinsky, T., He, C. J., Mitsuhashi, T., Bucala, R., Liu, C., Buenting, C., et al. (1997). Orally absorbed reactive glycation products (glycotoxins): An environmental risk factor in diabetic nephropathy. Proceedings of the National Academy of Sciences of the United States of America, 94, 6474–6479. Miyata, T., Taneda, S., Kawai, R., Ueda, Y., Horiuchi, S., Hara, M., et al. (1996). Identification of pentosidine as a native structure for advanced glycation end products in b2-microglobulin-containing amyloid fibrils in patients with dialysis-related amyloidosis. Proceedings of the National Academy of Sciences of the United States of America, 93, 2353–2358. Ohara-Takada, A., Matsuura-Endo, C., Chuda, Y., Ono, H., Yada, H., Yoshida, M., et al. (2005). Change in content of sugars and free amino acids in potato tubers under short-term storage at low temperature and the effect on acrylamide level after frying. Bioscience Biotechnology and Biochemistry, 69, 1232–1238. Pantazi, D., Papavergou, A., Pournis, N., Kontominas, M. G., & Savvaidis, I. N. (2008). Shelf-life of chilled fresh Mediterranean swordfish (Xiphias gladius) stored under various packaging conditions: microbiological, biochemical and sensory attributes. Food Microbiology, 25, 136–143. Riviere, S., Birlouez-Aragon, I., & Vellas, B. (1998). Plasma protein glycation in Alzheimer’s disease. Glycoconjugate Journal, 15, 1039–1042. Schmedes, A., & Hølmer, G. (1989). A new thiobarbituric acid (TBA) method for determination of free malonaldehyde (MDA) and hydroperoxides selectively as a measure of lipid peroxidation. Journal of the American Oil Chemists’ Society, 66, 813–817. Uribarri, J., Cai, W., Peppa, M., Goodman, S., Ferrucci, L., Striker, G., et al. (2007). Circulating glycotoxins and dietary advanced glycation endproducts: Two links to inflammatory response, oxidative stress, and aging. Journals of Gerontology Series A, Biological Sciences and Medical Sciences, 62, 427–433. Vlassara, H., Cai, W., Crandall, J., Goldberg, T., Oberstein, R., Dardaine, V., et al. (2002). Inflammatory markers are induced by dietary glycotoxins: A pathway for accelerated atherosclerosis in diabetes. Proceedings of the National Academy of Sciences of the United States of America, 99, 15596–15601. Wijewickreme, A. N., & Kitts, D. D. (1998). Metal chelating and antioxidant activity of model Maillard reaction products. Advances in Experimental Medicine and Biology, 434, 245–254. Yaylayan, V. A., & Huyghues-Despointes, A. (1994). Chemistry of Amadori rearrangement products: Analysis, synthesis, kinetics, reactions, and spectroscopic properties. Critical Reviews in Food Science and Nutrition, 34, 321–369. Zamora, R., & Hidalgo, F. J. (2005). Coordinate contribution of lipid oxidation and Maillard reaction to the nonenzymatic food browning. Critical Reviews in Food Science and Nutrition, 45, 49–59.
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Food Chemistry journal homepage: www.elsevier.com/locate/foodchem
Analytical Methods
Analysis of glycative products in sauces and sauce-treated foods Pei-chun Chao a,b, Cheng-chin Hsu b, Mei-chin Yin c,* a b c
Department of Nutrition, Chung Shan Medical University Hospital, Taiwan, ROC Department of Nutritional Science, Chung Shan Medical University, Taiwan, ROC Department of Nutrition, China Medical University, 91, Hsueh-shih Road, Taichung City, Taiwan, ROC
a r t i c l e
i n f o
Article history: Received 13 February 2008 Received in revised form 27 June 2008 Accepted 30 June 2008
Keywords: Pentosidine Carboxymethyllysine Soybean sauce Barbecue sauce Maillard reaction
a b s t r a c t Content of Maillard reaction products (MRPs) such as pentosidine, carboxymethyllysine and furosine in soybean sauce, sour-sweet sauce, tomato sauce, barbecue sauce, and sauce-treated chicken, pork, beef, salmon and cod was analysed. In test sauces, MRP content was in the range of 10–692 lg/100 mL sample. MRP content in raw, boiled, fried and baked foods was in the range of 10–76 lg/100 g sample. Boiling, frying and baking caused significantly higher MRP levels in test foods (P < 0.05). MRP levels in soybean sauce, sour-sweet sauce and barbecue sauce-treated foods were in the range of 1094–2424, 1494– 3146 and 1400–2926 lg/100 g sample. The interactions of sauce, heating and frying oil markedly enhanced the formation of MRPs in sauce-treated foods. Because MRPs are glycative products; thus, patients with glycation associated diseases may consider limiting the dietary use of these sauces. Ó 2008 Elsevier Ltd. All rights reserved.
1. Introduction Maillard reaction (MR) occurs between the free amino group of amino acids and the carbonyl group of reducing sugars such as glucose and fructose, and leads to the formation of reversible Amadori products, which spontaneously undergo further nonenzymatic rearrangement to form irreversible crosslinks called Maillard reaction products (MRPs) or advanced glycation endproducts (AGE) (Grandhee & Monnier, 1991; Yaylayan & Huyghues-Despointes, 1994). The common Amadori products or MRPs include pentosidine, carboxymethyllysine (CML) and furosine (Erbersdobler & Somoza, 2007; Goldberg et al., 2004). The presence of these MRPs in food may benefit food preservation because these MRPs could provide antioxidative protection (Bedinghaus & Ockerman, 1995). However, the impact of dietary MRPs upon healthy risk have been considered because dietary MRP content contributes to excess serum AGE levels, and the elevated AGE level in circulation favours the development of glycation and inflammation associated diseases such as renal failure, diabetes and Alzheimer’s disease (Koschinsky et al., 1997; Riviere, Birlouez-Aragon, & Vellas, 1998; Vlassara et al., 2002). Therefore, in order to avoid the adverse effects of dietary MRPs upon physiological variation of AGE levels, and the progression or deterioration of glycative injury, it is important to characterise and quantify MRPs in the diet. Soybean sauce, sour-sweet sauce, barbecue sauce and tomato sauce (ketchup) are less expensive and easily available sauces in * Corresponding author. Tel.: +886 4 22053366x7510; fax: +886 4 22062891. E-mail address: [email protected] (M.-c. Yin). 0308-8146/$ - see front matter Ó 2008 Elsevier Ltd. All rights reserved. doi:10.1016/j.foodchem.2008.06.076
Taiwan and Asia. Soybean sauce is a fermentative product of soybean, rich in protein. Sour-sweet sauce and barbecue sauce have sweet taste because sugar is an ingredient in these sauces. Therefore, these sauces are prone to undergo MR with foods rich in protein during food preparation and storage. Although these sauces are used for seasoning, they are used frequently and in a large amount in some food preparations in Asia. Thus, the influence of these sauces upon consumer’s healthy risk could not be ignored. So far, less information is available regarding the MRP content in these sauces, and sauce-treated foods. This study analysed the content of MRPs such as pentosidine, CML and furosine in commercially available soybean sauce, soursweet sauce, tomato sauce and barbecue sauce as well as foods rich in protein such as chicken, pork, beef, salmon and cod. The influence of boiling, frying or baking treatment upon MRP formation in foods and sauce-treated foods was also determined. The results of this study could provide novel information for people with glycation associated diseases to make appropriate dietary choices. 2. Materials and methods 2.1. Sauce and food sample Soybean sauce, sour-sweet sauce, tomato sauce (ketchup), barbecue sauce, chicken, pork, beef, salmon, and cod were purchased from local supermarkets in Taichung City (Taiwan). For commercial sauces, four brands of each sauce were examined, in which eight samples for each brand were used. Skin and visible fat of chicken, pork, beef, salmon, and cod were removed.
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2.2. Lipid oxidation measurement Food freshness could be evaluated by the level of lipid oxidation (Pantazi, Papavergou, Pournis, Kontominas, & Savvaidis, 2008). Thus, in order to ensure the freshness of chicken, pork, beef, salmon, and cod, lipid oxidation level of these food samples was determined by a thiobarbituric acid (TBA)-assay described by Schmedes and Hølmer (1989), in which 10 g sample were chopped and mixed with 25 mL 20% trichloroacetic acid solution (200 g/L of trichloroacetic acid in 135 mL/L phosphoric acid solution) and homogenised in a Waring Blender for 30 s. After filtration, a 2mL filtrate was combined with 2 mL, 0.02 M aqueous TBA in a test tube. The tubes were incubated at room temperature in the dark for 20 h, and the absorbance at 532 nm was measured by using UV–VIS spectrophotometer (model UV-1210, Shimadzu, Tokyo, Japan). Thiobarbituric acid reactive substances (TBARS) content were expressed as malondialdehyde equivalents (mg/kg food sample). In this study, malondialdehyde equivalents of used samples were in the range of 0.06–0.091 mg/kg food sample.
CML by reverse phase HPLC. Pentosidine was analysed by an HPLC method described by Miyata et al. (1996), in which a Supelco reverse phase C18 column (id, 4.6 mm; od, 25 cm; Supelco, Bellefonte, PA, USA) were equipped. The effluent was monitored by a fluorescence detector (RF-10A Shimadzu, Kyoto, Japan), and excitation and emission wavelength were set at 335 and 385 nm, respectively. The content of CML and pentosidine in used soybean oil, water and sauce for soaking, boiling or frying were analysed. CML and pentosidine levels in these samples were lower than 10 lg/100 mL. 2.6. Determination of furosine
The content of reducing sugar (glucose and fructose) in sauces and food samples was determined according to the method described by Ohara-Takada et al. (2005). Briefly, 10 mL sauce or 10 g food sample were mixed with 50 mL 80% ethanol. After 1 h heating at 80 °C, sample was cooled down to room temperature, and followed by centrifugation (2500g for 15 min at 25 °C). Supernatant was collected, dried under vacuum, re-dissolved in distilled water and passed through a 0.2-lm membrane filter. The concentration of glucose and fructose in filtrate was analysed by an HPLC (Shimadzu LC-10, Kyoto, Japan) quipped with an evaporative light scattering detector (Sedex, Paris, France) and a TSK gel Amide-80 column (id, 4.6 mm; od, 25 cm; Anachem, Luton, UK).
Furosine was determined according to the method described by Delgado, Corzo, Santa-Maria, Jimeno, and Olano (1992). Food or sauce-treated food, 50 g, was homogenised in 50 mL sterile distilled water. After filtrating through a Whatman No. 1 filter paper, the filtrate was collected. Then, 50 mL sauce or food filtrate were hydrolysed, using 3 mL of 7.95 M HCl at 110 °C for 23 h in screwcap tubes. Hydrolysis tubes were sealed under nitrogen. Hydrolysates were aerated and cooled down, and followed by centrifugation at 14,000g for 10 min. The supernatant was applied to a SepPak C18 cartridge (Millipore, Milfold, MA, USA) pre-wetted with 5 mL methanol and 10 mL deionized water, and followed by eluting with 3 mL 3 M HCl. The dried sample was re-dissolved in 1 mL mixture containing water, acetonitrile and formic acid at the ratio of 95:5:0.2 (v/v/v). Degassed mobile phase was prepared with 5 mM sodium heptane sulphonate including 20% acetonitrile and 0.2% formic acid. Furosine was quantified by the external standard method. Calibration curve was built from a stock solution in the range of 1–2000 lg/100 mL. The content of furosine in used soybean oil, water and sauce for soaking, boiling or frying were analysed. Furosine level in these samples was lower than 14 lg/ 100 mL.
2.4. Food preparation
2.7. Statistical analysis
In Asia, tomato sauce is not commonly used for cooking; thus, this sauce was not used to treat any food in this study. Each sauce used for food treatment was mixed from four commercial brands at equal amounts. Foods and sauce-treated foods were prepared as follows. For boiling preparation, 200 g food were cut into 4 strips and mixed with 400 mL sauce (or water), and boiled at 100 °C for 15 min. For deep frying preparation, 200 g food were cut into 4 strips and soaked with 400 mL sauce (or water) for 2 h at 25 °C. Then, sauce (or water) residue on food surface was removed by paper towel, and food sample was treated by deep frying in 100 mL soybean oil at 180 °C for 15 min. For baking preparation, 200 g food were cut into 4 strips and soaked with 400 mL sauce (or water) for 2 h at 25 °C. Then, sauce (or water) residue on food surface was removed by paper towel, and food sample was treated by baking at 230 °C for 15 min. After prepared, food was put on a Whatman No. 1 filter paper for 30 min for cooling and removing the residue of sauce or frying oil.
The effect of each treatment was analysed from eight different preparations (n = 8). Data were reported as means ± standard deviation (SD), and subjected to analysis of variance (ANOVA). Differences among means were determined by the least significance difference test with significance defined at P < 0.05.
2.3. Measurement of reducing sugars
2.5. Determination of carboxymethyllysine (CML) and pentosidine The method described by Inagi et al. (2006) was used to determine CML level. Sauce sample, 50 mL, or food and sauce-treated food sample, 50 g, was mixed with excess of NaBH4 in 0.2 mol/L borate buffer (pH 9.1) for reduction. Proteins were precipitated by 20% trichloroacetic acid, and centrifugation at 2000g for 10 min. The pellet was washed with 1 mL 10% trichloroacetic acid. After drying, the pellet was acid hydrolysed in 500 lL 6 M HCl for 16 h at 110 °C in screw-cap tubes purged with nitrogen. Hydrolysates were dried, and rehydrated in water, and used for measuring
3. Results Content of protein, lysine, glucose and fructose in soybean sauce, sour-sweet sauce, tomato sauce, barbecue sauce and food
Table 1 Content of protein (g), lysine (mg), glucose (mg) and fructose (mg) in 100 mL or 100 g soybean sauce, sour-sweet sauce, tomato sauce, barbecue sauce, chicken, pork, beef, salmon and cod Protein
Lysine
Glucose
Fructose
Soybean sauce Sour-sweet sauce Tomato sauce Barbecue sauce
5.2 1.6 1.2 3.6
204 –e – 178
16 ± 4a 37 ± 7c 10 ± 5a 23 ± 8b
7 ± 2a 58 ± 8c 41 ± 6b 76 ± 11d
Chicken Pork Beef Salmon Cod
13.4 12.4 9.8 13.0 8.8
1452 1868 1728 1626 1540
– – – – –
– – – – –
Data are mean ± SD, n = 8. Protein and lysine data are from Taiwan Nutrient Database. a–d Means in a column without a common letter differ, P < 0.05. e Means too low to be detected.
4. Discussion Our present study found 4 sauces contained furosine, CML and pentosidine. Furosine is a degradation product of Amadori compounds; CML and pentosidine are irreversible crosslinks formed from Amadori products during advanced stages of Maillard reactions. Therefore, the presence of these three MRPs strongly implied that Maillard reaction has occurred, and might be still undergoing in these sauces. Furthermore, we found the content of pentosidine, CML and furosine in each type of sauce was from brand to brand. It is known that the MRPs content in foods is related to their composition, the method for processing and condition for storage (Chuyen, 2006; Goldberg et al., 2004). Because these sauces contained various levels of protein, lysine and reducing sugars (glucose and
Table 2 Content (lg/100 mL sample) of pentosidine, CML and furosine in soybean sauce (SB), sour-sweet sauce (SS), tomato sauce (TS) and barbecue sauce (BB) Sauce
MRP
Range
Mean (n = 8)
SB
Pentosidine CML Furosine
10–82 172–392 28–102
42 ± 18 224 ± 43 58 ± 26
SS
Pentosidine CML Furosine
24–104 124–532 24–110
66 ± 25 306 ± 63 74 ± 19
TS
Pentosidine CML Furosine
20–76 176–692 42–120
52 ± 16 416 ± 89 60 ± 21
BB
Pentosidine CML Furosine
78–158 252–482 26–110
104 ± 46 352 ± 70 76 ± 17
90
d
70
d
d
d
60
raw
c c
c c
50
c
b
boiled
b
b
40
fried
b
30 20
e
e
80
a
baked
a
a
a
a
10 0 chicken
pork
90
CML (ug/100 g sample)
samples is shown in Table 1. Food samples had more protein and lysine than sauces; however, sauces had more glucose and fructose than food samples. The content of glucose in sauces followed the order: sour-sweet sauce > barbecue sauce > soybean sauce = tomato sauce (P < 0.05). Fructose content in sauces followed the order: barbecue sauce > sour-sweet sauce > tomato sauce > soybean sauce (P < 0.05). The content of pentosidine, CML and furosine in soybean sauce, sour-sweet sauce, tomato sauce, and barbecue sauce is presented in Table 2. The contents of these three MRPs were different from brand to brand, and in the range of 10– 692 lg/100 mL sauce. The content of pentosidine, CML and furosine in raw, boiled, fried or baked chicken, pork, beef, salmon and cod is shown in Fig. 1. In test foods, pentosidine, CML and furosine contents were in the range of 10–72, 14–76, and 22–68 lg/ 100 g food. The process of boiling, frying and baking significantly increased levels of pentosidine, CML and furosine in test foods (P < 0.05). In salmon and cod, frying process caused significantly higher levels of pentosidine and CML than boiling or baking process (P < 0.05). The content of pentosidine, CML, furosine and sum of these three glycative products in boiled, fried and baked chicken, pork, beef, salmon and cod treated by soybean sauce, sour-sweet sauce or barbecue sauce is shown in Tables 3–5, respectively. The sum of pentosidine, CML and furosine levels in soybean sauce-treated foods were in the range of 1094–2424 lg/100 g sample. Soursweet sauce-treated foods had total level of pentosidine, CML and furosine in the range of 1494–3146 lg/100 g sample. Barbecue sauce-treated foods had total level of pentosidine, CML and furosine in the range of 1400–2926 lg/100 g sample. For each test food sample, frying and baking treatments caused significantly greater formation of pentosidine, CML and furosine than boiling treatment (P < 0.05).
pentosidine (ug/100 g sample)
P.-c. Chao et al. / Food Chemistry 113 (2009) 262–266
beef
salmon
cod
d
d
d
d
80 70
c
c
c
c c
c
60 50
b
b
b
b
b
boiled fried
40
baked
a
30 20
raw
a
a a
a
10 0 chicken
pork
80
furosine (ug/100 g sample)
264
beef
salmon
cod
d
d d
70 c 60
c
c
c
c
c
c c c
c
b
50
b
40 a
fried
a
30
raw boiled
a a
a
baked
20 10 0 chicken
pork
beef
salmon
cod
Fig. 1. Content (lg/100 g sample) of pentosidine, CML and furosine in raw, boiled, fried or baked chicken, pork, beef, salmon and cod. Data are mean ± SD (n = 8). (a–e) Means among bars without a common letter differ, P < 0.05.
Table 3 Content (lg/100 g sample) of pentosidine, CML and furosine in boiled, fried and baked chicken, pork, beef, salmon and cod treated by soybean sauce Sample
Method
Pentosidine
CML
Furosine
Sum
Chicken
Boiled Fried Baked
192 ± 24 206 ± 29 254 ± 23
638 ± 45 878 ± 48 694 ± 41
688 ± 49 752 ± 40 900 ± 47
1518 ± 52c 1836 ± 68d 1848 ± 74d
Pork
Boiled Fried Baked
174 ± 20 260 ± 25 324 ± 30
572 ± 48 904 ± 44 922 ± 60
840 ± 51 692 ± 38 962 ± 63
1526 ± 66c 1856 ± 60d 2208 ± 87e
Beef
Boiled Fried Baked
182 ± 18 291 ± 31 340 ± 27
526 ± 49 854 ± 61 764 ± 42
820 ± 40 654 ± 25 776 ± 43
1538 ± 69c 1798 ± 80d 1880 ± 93d
Salmon
Boiled Fried Baked
148 ± 20 238 ± 29 256 ± 31
486 ± 47 980 ± 50 1078 ± 65
460 ± 32 1206 ± 72 954 ± 59
1094 ± 55a 2424 ± 84f 228 ± 83e
Cod
Boiled Fried Baked
160 ± 19 218 ± 33 270 ± 27
550 ± 58 846 ± 76 952 ± 64
544 ± 44 858 ± 62 688 ± 50
1254 ± 71b 1922 ± 89d 1910 ± 73d
Data are mean ± SD, n = 8. a–f Means in a column without a common letter differ, P < 0.05.
fructose); thus, MRPs could be formed naturally in these sauces during storage. In addition, the results of our present study revealed that fried salmon and cod had higher pentosidine and CML levels than baked samples, although the baking temperature (230 °C) was higher than frying temperature (180 °C). It was notified that heat directly acted on food during the baking process; however, during frying process, heat acted on frying oil first and food obtained heat from the hot frying oil. Thus, MRPs presented in fried food may comprise
P.-c. Chao et al. / Food Chemistry 113 (2009) 262–266 Table 4 Content (lg/100 g sample) of pentosidine, CML and furosine in boiled, fried and baked chicken, pork, beef, salmon and cod treated by sour-sweet sauce Sample
Method
Pentosidine
CML
Furosine
Sum
Chicken
Boiled Fried Baked
152 ± 20 190 ± 31 334 ± 33
906 ± 57 1350 ± 76 1486 ± 81
512 ± 38 462 ± 34 680 ± 47
1570 ± 62a 2002 ± 75c 2500 ± 90c
Pork
Boiled Fried Baked
164 ± 23 226 ± 36 310 ± 29
808 ± 66 1242 ± 79 1426 ± 91
522 ± 45 668 ± 56 604 ± 53
1494 ± 73a 2136 ± 83c 2340 ± 104d
Beef
Boiled Fried Baked
186 ± 27 254 ± 42 362 ± 40
870 ± 71 1364 ± 83 1208 ± 68
527 ± 32 790 ± 60 684 ± 55
1578 ± 77a 2408 ± 92e 2254 ± 73d
Salmon
Boiled Fried Baked
170 ± 25 302 ± 29 276 ± 34
1084 ± 59 1670 ± 86 2180 ± 101
528 ± 34 924 ± 56 690 ± 49
1782 ± 70b 2896 ± 92g 3146 ± 120h
Cod
Boiled Fried Baked
154 ± 19 188 ± 32 258 ± 39
974 ± 78 1522 ± 89 1924 ± 91
612 ± 59 1064 ± 88 766 ± 76
1740 ± 92b 2774 ± 95f 2948 ± 112g
Data are mean ± SD, n = 8. a–h Means in a column without a common letter differ, P < 0.05.
Table 5 Content (lg/100 g sample) of pentosidine, CML and furosine in boiled, fried and baked chicken, pork, beef, salmon and cod treated by barbecue sauce Sample
Method
Pentosidine
CML
Furosine
Sum
Chicken
Boiled Fried Baked
452 ± 46 490 ± 56 634 ± 43
542 ± 47 844 ± 63 1152 ± 87
406 ± 45 568 ± 60 778 ± 66
1400 ± 73a 1902 ± 82c 2564 ± 115f
Pork
Boiled Fried Baked
460 ± 38 534 ± 57 602 ± 64
620 ± 36 882 ± 65 1226 ± 71
680 ± 60 736 ± 65 646 ± 54
1760 ± 74b 2152 ± 90d 2474 ± 97f
Beef
Boiled Fried Baked
486 ± 55 562 ± 58 672 ± 67
588 ± 69 790 ± 78 1168 ± 95
456 ± 38 534 ± 50 650 ± 63
1530 ± 74a 1886 ± 82c 2490 ± 103f
Salmon
Boiled Fried Baked
440 ± 46 614 ± 70 546 ± 58
632 ± 62 1030 ± 92 1522 ± 106
618 ± 65 1144 ± 85 858 ± 60
1690 ± 76b 2788 ± 94g 2926 ± 88h
Cod
Boiled Fried Baked
434 ± 35 508 ± 49 528 ± 61
546 ± 57 828 ± 86 1506 ± 92
522 ± 43 980 ± 76 704 ± 54
1502 ± 70a 2316 ± 91e 2738 ± 108g
Data are mean ± SD, n = 8. a–h Means in a column without a common letter differ, P < 0.05.
of MRPs from both heat-treated frying oil and frying oil-treated food. We also analysed the content of CML and pentosidine in used soybean oil and found that CML and pentosidine levels in these samples were lower than 10 lg/100 mL. Thus, the MRPs presented in fried foods in our present study were mainly from the interaction of hot frying oil and food. Soybean oil was used in this study for frying treatment. This oil and two test fish are rich in unsaturated fatty acids. It has been reported that MRPs could be formed from Schiff base and Amadori adducts by autoxidation of fatty acids and amino acids (Bucala, Makita, Koschinsky, Cerami, & Vlassara, 1993; Fu et al., 1996). Zamora and Hidalgo (2005) also indicated that lipid oxidation and Maillard reaction were interrelated, and lipid oxidation products could affect Maillard pathway. Thus, it is possible that heat caused oxidation of unsaturated fatty acids from soybean oil and fish, and the oxidative products participated the rearrangement of Amadori adducts to form irreversible crosslinks like pentosidine or CML. Furthermore, we also notified that fried or baked sauce-treated salmon had markedly higher sum of CML, pentosidine and furosine than fried or baked sauce-treated cod. Salmon has 54.9% monounsaturated fatty acids (MUFA) and 24.3% polyunsaturated fatty acids (PUFA); how-
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ever, cod has 73.1% MUFA and 6.1% PUFA (Department of Health, 1998). Obviously, the role of PUFA on MRP formation can not be ignored. These findings suggest that frying or baking treatment should be limited for foods rich in unsaturated fatty acids in order to reduce MRPs formation in foods. Furosine is a direct marker of lysine reaction product and also a relevance for nutritionally unavailable for lysine from food source (Bujard & Finot, 1978; Del Castillo, Corzo, & Olano, 1999). Lysine content of test foods in our present study was in the range of 1452–1868 mg/100 g (Department of Health, 1998). Our present study found the furosine levels of sauce and foods treated by boiling, frying or baking were in the range of 24–120 and 24–68 lg/ 100 mL or 100 g, respectively. However, furosine level in saucetreated foods was in the range of 406 (boiled barbecue sauce-treated chicken)–1206 (fried soybean sauce-treated salmon) lg/100 g. Obviously, about 200–1000 lg/100 g newly produced furosine resulted from MR during food preparation and processing, in which lysine from foods was involved. After calculation, we found that at least 100–540 lg lysine/100 g sauce-treated foods (equal to 0.8–4.5 mg lysine/100 g protein) was nutritional unavailable. Pentosidine could be formed from arginine and lysine (Grandhee & Monnier, 1991). Thus, the formation of pentosidine in sauce-treated foods as we observed also indicated not only thermal damage in food protein but also loss of nutritional available arginine or lysine. Fu et al. (1996) reported that CML could be produced by many pathways such as degradation of the Amadori compounds or direct reaction of lysine with some sugars or lipid oxidation derived dicarbonyl compounds. Erbersdobler and Somoza (2007) indicated that in more severely heat-treated food items, in which furosine levels have already decreased, CML can provide additional information on the protein damage. In our present study, the CML level in baked sour-sweet sauce-treated salmon and cod was very high (2180 and 1924 lg/100 g sample). This finding revealed that advanced MR had occurred in these food samples and also implied that these foods had been severely heat-treated, in which baking treatment provided heat, sour-sweet sauce provided reducing sugars, salmon (or cod) provided lysine and/or unsaturated fatty acids. Thus, CML formation in these food samples involved many factors and complicated chemical reactions. Therefore, in order to maintain protein quality of food and decrease the production of MRPs including CML, pentosidine and furosine, it is better to reduce using baking process and/or sauces for food preparation. On the other hand, we found sour-sweet sauce contained more reducing sugars than soybean sauce, and sour-sweet sauce-treated foods had more MRPs than soybean sauce-treated foods. This finding agreed that reducing sugar is an important factor contributed to MRPs formation, and also partially explains the impact of sauce upon MRP formation in sauce-treated foods. In addition, pentosidine (or CML) level in each sauce-treated food was higher than the sum of pentosidine (or CML) from corresponding sauce, and pentosidine (or CML) from corresponding food. For instance, the CML level of soybean sauce and boiled chicken were 224 lg/ 100 mL and 30 lg/100 g, respectively. However, the CML level in boiled soybean sauce-treated chicken was 638 lg/100 g. Also, the pentosidine level of barbecue sauce and fried salmon was 104 lg/100 mL and 70 lg/100 g only; but, the pentosidine level of fried barbecue sauce-treated salmon was 614 lg/100 g. These results revealed that sauce and heat caused a synergistic effect on MRPs formation in these sauce-treated foods. It is highly possible that heat caused more amino acids release from the tested foods, which allowed reducing sugars from sauces to react with the released amino acids and facilitated the formation of MRPs. Tomato sauce, or so-called ketchup, is commonly used for foods popular in western society such as French fried, hamburger and hot dog. Soybean sauce, sour-sweet sauce, and barbecue sauce are popular in Asia for food preparation. Although they are commonly used
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for seasoning only, they are used with high frequency and large quantity in certain food preparations. In Taiwan and Japan, soybean sauce could be directly used as table sauce like ketchup for foods such as tofu and sashimi (raw fish). It has been indicated that MRPs possess antioxidative property and could extend shelf-life of food via delaying lipid oxidation (Bedinghaus & Ockerman, 1995; Wijewickreme & Kitts, 1998); thus, these MRPs might benefit food preservation. However, the dietary MRPs could elevate AGE levels in circulation, which consequently contributes to the progression of glycation associated diseases such as diabetes, Alzheimer’s disease and inflammatory arthritis (Uribarri et al., 2007). Thus, patients with glycation associated diseases should avoid consuming foods or sauces rich in MRPs based on healthy consideration. In conclusion, soybean sauce, sour-sweet sauce, tomato sauce, barbecue sauce contained reducing sugars, and the content of pentosidine, CML and furosine in these sauces was in the range of 10–692 lg/100 mL. MRP content in sauce-treated foods was in the range of 1094–3146 lg/100 g. Besides heat, frying oil played a role to affect the formation of MRPs in test foods. Furthermore, sauce and heat caused a synergistic effect in the formation of MRPs in sauce-treated foods. These data suggest that patients with glycation associated diseases should consider limiting the dietary use of these sauces. References Bedinghaus, A. J., & Ockerman, H. W. (1995). Antioxidative Maillard reaction products from reducing sugars and free amino acids in cooked ground pork patties. Journal of Food Science, 60, 992–995. Bucala, R., Makita, Z., Koschinsky, T., Cerami, A., & Vlassara, H. (1993). Lipid advanced glycosylation: Pathway for lipid oxidation in vivo. Proceedings of the National Academy of Sciences of the United States of America, 90, 6434–6438. Bujard, E., & Finot, P. A. (1978). Measure of available and blocked lysine in industrial milks. Annales de la nutrition et de l’alimentation, 32, 291–305. Chuyen, N. V. (2006). Toxicity of the AGEs generated from the Maillard reaction: On the relationship of food-AGEs and biological-AGEs. Molecular Nutrition and Food Research, 50, 1140–1149. Del Castillo, D. M., Corzo, N., & Olano, A. (1999). Early stages of Maillard reaction in dehydrated orange juice. Journal of Agricultural and Food Chemistry, 47, 4388–4390. Delgado, T., Corzo, N., Santa-Maria, G., Jimeno, M. L., & Olano, A. (1992). Determination of furosine in milk samples by ion pair reversed phase liquid chromatography. Chromatographia, 33, 374–376. Department of Health (1998). Taiwan nutrient databases 1998. Taipei, Taiwan: Department of Health, Executive Yuan. Erbersdobler, H. F., & Somoza, V. (2007). Forty years of furosine – forty years of using Maillard reaction products as indicators of the nutritional quality of foods. Molecular Nutrition and Food Research, 51, 423–430.
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