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Journal of Food Science and Technology logoLink to Journal of Food Science and Technology
. 2020 Sep 24;58(8):3001–3009. doi: 10.1007/s13197-020-04803-w

Effect of basil use in meatball production on heterocyclic aromatic amine formation

Idil Uzun 1, Fatih Oz 1,✉
PMCID: PMC8249626  PMID: 34294962

Abstract

Herein, the effects of basil usage in meatball production on various quality criteria and heterocyclic aromatic amine (HAA) formation were investigated. The use of basil at every rate caused a significant reduction in TBARS value compared to control group. Cooking caused an increase in pH and TBARS values. IQx, IQ, AαC and MeAαC compounds could not be detected, while various amounts of MeIQx, MeIQ, 7,8-DiMeIQx, 4,8-DiMeIQx and PhIP were determined in the samples. Total HAA contents were determined up to 1.61 ng g−1 and increasing of cooking temperature increased total HAA content, except for meatball with 1% basil. The reducing or enhancing effect of the use of basil in meatball production on the formation of HAA varied depending on the usage rate and cooking temperature. It was determined that even if 100 g of the meatballs containing 0.5% basil cooked at 250 °C whose total amount of HAA content was the highest, is eaten, the intake amount is far below 1 μg.

Keywords: Meatball, Sweet basil, Cooking, Heterocyclic aromatic amine, TBARS

Introduction

Foods are a complex mixture of macro components (protein, fat, and carbohydrate) and micro components (vitamins and minerals) that have many important functions. In addition, antioxidants and dietary fibers are also found in foods and have an important role in regulating human nutrition. However, foods may also contain many harmful components such as unwanted chemicals, microbial contaminants, and various toxins. For these reasons, foods that play an important role in the prevention or treatment of some diseases are also responsible for the emergence of some diseases (Oz and Kaya 2011a).

Meat, an essential part of the human diet, is usually subjected to heat treatment just before its consumption. Cooking meat is necessary to obtain a delicious and safe product. However, due to the heat treatment applied, undesirable changes such as loss of vitamins and minerals, changes in fatty acid composition due to lipid oxidation, and decrease in nutritional value may occur (Rodriguez-Estrada et al. 1997; Gerber et al., 2009). Various food toxicants may also form in meat (Oz and Kaya 2011a). In 1977, it was first reported that mutagenic substances could be formed as a result of cooking the protein-rich foods. Among these compound, HAAs are very important (Oz and Kaya 2011a).

HAAs are mutagenic and/carcinogenic compounds that are formed in meat and fish products cooked at generally 150 °C or over (Oz and Kaya 2011a). Until today, more than 25 HAAs have been isolated and identified in foods (Oz 2019) and defined as strong mutagens (Oz 2020). It is known that the International Agency for Research on Cancer (IARC) has classified some HAAs as “possible human carcinogens” and “probable human carcinogens” (Oz 2020). HAA type and amounts formed in foods depend on various factors such as meat type, cooking procedures, pH, water activity, carbohydrates, free amino acids, creatine, heat and mass transfer, fats, lipid oxidation and antioxidant (Jägerstad et al. 1998; Oz 2020).

Exposure to HAAs has been associated with many different types of cancer and mutagenesis. Therefore, the prevention of HAA formation during food processing procedures is attracting attention. Because free radicals are involved in the HAA formation mechanism, antioxidants are considered to be the most promising and effective inhibitors of the HAA formation mechanism. Spices that are commonly used in meat processing are an important source of antioxidants (Zeng et al. 2017).

Basil, Ocimum basilicum L., is an economically important plant and an aromatic spice, belonging to Lamiaceae family, which has bright green leaves and can reach 90 cm in height. It is used in kitchens due to its pleasant smell and taste. Although it is an Asian, African, Central and South American plant, it is widely cultivated in many countries, especially in the Mediterranean region. Its essential oils are used as a flavoring agent in the food and beverage industry, a fragrance in pharmaceutical and industrial products and a fungicide and insecticide (Copetta et al. 2006). Basil is also called "the king of plants" due to its intense aroma, potent antioxidant capacity, antimicrobial activity and anticarcinogenic activity. These properties of basil are attributed to the content of its phenolic and aromatic substances (Lee and Scagel 2009; Filip 2017).

Antioxidants are the substances that delay the oxidation of easily oxidizable biomolecules such as lipids and proteins in foods, thus protecting the food against spoilage caused by oxidation, and extend the shelf life of the food (Karre et al. 2013). Especially due to the potential carcinogen activity of synthetic antioxidants, there is much more interest in the antioxidant activity of natural substances in recent years (Jayasinghe et al. 2003). Many studies have reported the effects of natural (flavonoids, vitamins E and C) and synthetic antioxidants [such as butylated hydroxy toluene (BHT), butylated hydroxy anisole (BHA), etc.], as well as some food components (cherry, a phenolic compound in tea, olive oil, spices) on the formation HAAs (Ahn and Grun 2005; Cheng et al. 2009; Oz and Cakmak 2016; Oz 2019). There are many studies in the literature showing the reducing and/or preventing effect of different spices used in the meatball preparation on the formation of HAAs. However, there is only one study (Damašius et al. 2011) on basil that has proven to have an antioxidant and anticarcinogenic effect. In that study, ethanol–water extract of basil was used and only the formation of a limited number of HAA compounds (PhIP, Trp-P-1, and Trp-P-2) was examined. On the other hand, although many studies have been conducted on the content of HAA in meatballs and the effects of some spices and plant extracts on the formation of HAAs, there are no studies examining the effects of different rates of basil usage on the meatball production on the formation of HAAs. Therefore, the present study was planned and carried out to determine the effect of different rates (0.25, 0.5, 0.75 and 1%, w/w) of basil usage in meatball production on the formation of HAAs (IQx, IQ, MeIQx, MeIQ, 7,8-DiMeIQx, 4,8-DiMeIQx, PhIP, AαC, and MeAαC) in meatballs cooked at different temperatures (150 °C, 200 °C and 250 °C). In addition, the effect of basil usage on the various quality criteria (water, pH, lipid oxidation and cooking loss) of the meatballs was also determined.

Materials and methods

Material

Raw material

In the present study, beef meat (M. Longissimus dorsi) was obtained from the Meat and Milk Institution (Erzurum, Turkey) and brought to the laboratory under the cold chain. Then it was separated from the visible fat and connective tissues and ground through a 3 mm plate grinder. The fat contents of the meatball were adjusted to 15% with the intermuscular fat from the same carcass. Basil was obtained in a packaged powder form from a super market in Erzurum (Turkey).

Chemicals

HAAs standards; 2-amino-3-methylimidazo[4,5-f]quinoxaline (IQx), 2-amino-3-methylimidazo[4,5-f]quinolone (IQ), 2-amino-3,8-dimethylimidazoimidazo[4,5-f]quinoxaline (MeIQx), 2-amino-3,4-dimethylimidazoimidazo[4,5-f]quinolone (MeIQ), 2-amino-3,7,8-trimethylimidazo[4,5-f]quinoxaline (7,8-DiMeIQx), 2-amino-3,4,8-trimethylimidazo[4,5-f]quinoxaline (4,8-DiMeIQx), 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), 2-amino-9H-pyrido[2,3-b]indole (AαC), 2-amino-3-methyl-9H-pyrido[2,3-b]indole (MeAαC) and 2-amino-3,4,7,8-tetramethylimidazo[4,5-f]quinoxaline (4,7,8-TriMeIQx) were purchased from Toronto Research Chemicals (Toronto, Canada). 4,7,8-TriMeIQx was used as the internal standard (Oz 2020).

Methods

The preparation of meatball dough

After the fat content of the minced meat was adjusted to 15% with the intermuscular fat, the meatball dough (2.5 kg) was divided into five portions (500 g), where one of these portions was used as the control group (without basil). Basil were directly added to the rest of the groups at four different rates (0.25, 0.5, 0.75 and 1%, w/w) and well mixed. Then, the meatball doughs stored at 4 °C for 6 h so as to allow the penetration of basil into the meatballs. The dough in each group was shaped into meatballs (1 cm thick and 80 mm diameter) using a metal shaper. The weight of the meatballs was about 70 g. No any additives were used to meatball dough to avoid any interaction.

Cooking conditions

As a result of preliminary tests, the meatballs were cooked on a hot plate preheated at 150 °C, 200 °C and 250 °C for 8 (4 + 4) min without adding frying fat or oil. Cooking temperature was measured by using a laboratory type thermometer (Part no: 0560 9260, Testo 926, Lenzkirch, Germany) and cooking time was measured by using a laboratory type chronometer. The internal temperatures of all of the meatballs were higher than 71.1 °C which is the recommended internal temperature for ground meats by the United States Department of Agriculture, Food Safety and Inspection Service (USDA–FSIS). After the meatballs cooled down to room temperature, the cooking loss was calculated, then the samples were homogenized with a kitchen type blender to have a uniform sample for the analyses. The analyses except for the HAA analysis were done on the same day. However, a little amount from the samples was vacuum packaged and stored at −18 °C for the HAA analysis. Freezing was done so that the samples could be stored before clean-up without any risk of further reactions in the meatballs. The frozen samples were thawed in a refrigerator at 4 °C for 12 h prior to use.

Water and pH analysis

Water contents and pH values of the samples were determined according to Gokalp et al. (2010).

Thiobarbituric acid reactive substances analysis

In the determination of lipid oxidation in the samples, the analysis of thiobarbituric acid reactive substances (TBARS) was used. TBARS value was determined by the method of Kılıç and Richards (2003). TBARS values of the samples were given as mg MDA kg−1 sample.

Cooking loss analysis

The weights of the meatball samples were measured at room temperature before and after cooking. Weight losses were calculated as % by determining the weights of the sample before and after cooking at room temperature.

Free radical scavenging activity (DPPH) analysis

The preparation of basil-water extract was done according to the method of Nuray and Oz (2019), while for the analysis of the free radical scavenging activity of the lyophilized basil-water extract, the method applied by Blois (1958) was used.

Heterocyclic aromatic amine analysis

The HAA analyses of the samples were done according to Oasis method. 1 g cooked sample and 12 ml 1 M NaOH in a beaker were mixed with a magnetic stirrer for 1 h and 10 g Extrelut NT packaging material NT (refill material, Merck, Darmstadt, Germany) was then added and mixed. The packaging process was done by using Oasis MCX cartridge (3 cm3, 60 mg, Waters, Milford, Massachusetts, USA) and Bond reservoir (Extrelut-20) in vacuum manifold system (Supelco, Visiprep). The HAAs were extracted with ethyl acetate, methanol, and hydrochloric acid, and finally, the extract was eluted with 2 ml MeOH:NH3 (19:1). Samples extracts were stored at −18 °C until HPLC analysis. One day prior to HPLC analysis, methanol and ammonia solutions were allowed to evaporate overnight in a drying oven at 45 °C, then 100 µl methanol with the internal standard was added and run in HPLC–DAD system (Oz 2020).

The amounts of HAAs were determined by using HPLC (Thermo Ultimate 3000, Thermo Scientific, USA) with diode array detector (DAD-3000). Separation process was performed at 35 °C and 0.7 ml min−1 flow rate on AcclaimTM 120 C18 3 mm (4.6 × 150 mm) Tosoh Bioscience GmbH (Stuttgart, Germany) by using methanol/acetonitrile/water/acetic acid (8/14/76/2, v/v/v/v, adjusted pH 5.0 with 25% ammonium hydroxide) as Solvent A and acetonitrile (100%) as Solvent B (Oz 2020). Gradient programme was set as 0% B, 0–10. min; 0–23% B, 11–19 min; 100% B, 20–22 min; 0% B, 23–33 min 10 µl injection was taken from each sample.

Statistical analysis

The present study was carried out according to a completely randomized design and employed in two replicates. The results were analyzed using SPSS package program and Duncan multiple comparison tests was used to evaluate the differences between the average values found.

Results and discussion

Water, pH and TBARS results of raw materials

The water content, pH and TBARS values of the beef meat (M. Longissimus dorsi), intermuscular fat obtained from the same carcass and raw meatballs with 15% fat content are shown in Table 1. Similar results were found by other researchers (Nuray and Oz 2019).

Table 1.

Water content, pH and TBARS values of beef, intermuscular fat and raw meatball with 15% fat

n Water (%) ± SD pH ± SD TBARS (mg MDA kg−1) ± SD
Meat 2 70.38 ± 0.55a 5.79 ± 0.01c 0.293 ± 0.079a
Intermuscular fat 2 19.78 ± 0.08c 6.77 ± 0.01a 0.251 ± 0.033a
Raw meatball 2 66.14 ± 0.98b 5.90 ± 0.01b 0.539 ± 0.401a
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Different letters (a–c) in the same column are significantly different (p < 0.05)

SD Standard deviation

Free radical scavenging activity of basil

In the present study, the free radical scavenging activity of basil used in the production of meatball and gallic acid used as a standard antioxidant were determined by using the DPPH method. It was determined that basil and gallic acid were not statistically different from each other in terms of DPPH free radical scavenging activities (p > 0.05).

Water content of the meatballs

The water contents of the samples are shown in Table 2. The use of basil at different rates in meatballs production had no statistically significant effect (p > 0.05) on the water content of meatballs. It is thought that the low dry matter content (9.05 ± 0.31%) of basil used in the production of meatballs has influenced this result. Indeed, Celebi (2010) reported that dry matter contents of basil samples of different companies varied between 8.42% and 11.41%. In parallel with the results of the present research, Teye et al. (2013) reported that basil usage in meatballs prepared by adding different amounts of basil (2, 4 and 6 g kg−1) had no significant effect on water content. Cooking, as expected, caused a statistically significant (p < 0.01) decrease in the water content of the samples and this situation is attributed to the shrinking observed in the myofibrillar proteins and perimysial connective tissue (del Pulgar et al. 2012). Similar results were found by other researchers (Oz and Cakmak 2016; Oz et al. 2016).

Table 2.

Effects of usage rate of basil, cooking process, and cooking temperatures on water content, pH, TBARS and cooking loss values of the samples

n Water (%) ± SD pH ± SD TBARS (mg MDA kg−1) ± SD Cooking loss (%) ± SD
Usage rate (UR, %)
 0 12 61.33 ± 4.36a 5.99 ± 0.14a 0.816 ± 0.338a 35.32 ± 3.61ab
 0.25 12 60.92 ± 4.76a 5.98 ± 0.13a 0.604 ± 0.066b 36.33 ± 4.33a
 0.5 12 61.08 ± 5.32a 5.99 ± 0.16a 0.588 ± 0.160b 35.58 ± 5.29ab
 0.75 12 61.00 ± 5.25a 5.98 ± 0.13a 0.633 ± 0.119b 33.83 ± 4.59bc
 1 12 60.77 ± 4.97a 5.98 ± 0.12a 0.659 ± 0.094b 32.77 ± 4.64c
 Sign NS NS ** *
Cooking process (CP)
 Raw 30 65.34 ± 0.80a 5.86 ± 0.01b 0.571 ± 0.088b
 Cooked 30 56.70 ± 2.68b 6.11 ± 0.05a 0.748 ± 0.231a
 Sign ** ** **
Cooking temperature (CT, °C)
 150 20 62.73 ± 2.79a 5.99 ± 0.14b 0.673 ± 0.273a 29.33 ± 2.00c
 200 20 60.35 ± 5.23b 5.96 ± 0.10c 0.650 ± 0.136a 36.69 ± 1.81b
 250 20 59.98 ± 5.57b 6.00 ± 0.15a 0.656 ± 0.158a 38.27 ± 2.14a
 Sign ** ** NS **
Interactions
 UR x CP NS ** **
 UR x CT NS * NS NS
 CP x CT ** ** NS
 UR x CP x CT NS * NS
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Different letters (a–c) in the same column are significantly different (p < 0.05)

SD Standard deviation, NS non-significant (p > 0.05), *: p < 0.05, **: p < 0.01

pH values of the meatballs

The pH values of the samples are also shown in Table 2. The use of different rates of basil in meatball production had no statistically significant effect (p > 0.05) on the pH values of the meatballs. The pH value of the basil used in meatball production was found to be 5.99 ± 0.01. Therefore, it is thought that the usage amounts are not at the level to affect the pH values of meatballs. Similarly, Celebi (2010) reported that pH values of basil samples of different companies ranged between 5.61 and 5.98. On the other hand, Teye et al. (2014) reported that pH values of pork meatballs prepared by adding basil significantly decreased. Cooking significantly increased pH values of the meatballs (p < 0.05). This situation is due to the fact that cooking causes the bonds containing imidazole, sulfhydryl and hydroxyl groups within meat to be released (Oz 2019). Similar results were determined by other researchers (Tengilimoglu-Metin et al. 2017).

TBARS results of the meatballs

Lipid oxidation is an important biochemical phenomenon that causes a great number of hydroperoxides production and leads to the formation of many different volatile compounds through different decomposition pathways (Jin et al. 2010). It is also an important factor in the development of sensory properties of the meat products (Harkouss et al. 2015). Gandemer (2002) reported that processing conditions affect the kinetics of oxidation reaction to a large extend. TBARS values of the samples are also shown in Table 2. The use of basil in the meatball production significantly decreased TBARS value compared to the control group without basil. On the other hand, there was no statistically significant difference (p > 0.05) between different basil usage rates (0.25, 0.5, 0.75 and 1%, w/w) in terms of TBARS values. This result shows that even 0.25% basil usage in meatball production was sufficient to reduce lipid oxidation. In parallel with the present research, Gulcin et al. (2007) declared that water and ethanol extracts of basil have very strong antioxidant activity. The antioxidant activity of basil has been connected to being rich in phenolic substances (Lee and Scagel 2009; Celebi 2010; Filip 2017). Indeed, Lee and Scagel (2009) found total polypheolic amount as 208 and 236 mg gallic acid 100 g−1 tissue in sweet basil and Thai basil, respectively. There are also other studies showing that lipid oxidation level of meat and meat products with basil is lower than control group samples without basil (Juntachote et al. 2006; Cichoski et al. 2011). Indeed, Juntachote et al. (2006) declared that TBARS value that was 2.04 mg MDA/kg in control group ground pork samples reduced to 1.22 mg MDA/kg as a result of using Holy basil extract. Cooking significantly increased TBARS value of the meatballs (p < 0.01). Similar results were determined by other researchers (Domínguez et al. 2014). Although it is known that many factors such as temperature, oxygen, light, catalysts etc. affect lipid oxidation in foods, it is also known that heat treatment affects on the level of lipid oxidation in meat and meat products. Therefore, it was expected that lipid oxidation in meat and meat products would increase by cooking. The reason of this is attributed to the iron released from myoglobin and hemoglobin denatured by the cooking process catalyzes the lipid oxidation (Nuray and Oz 2019). In addition, cooking meat destroys the cell structure and can cause prooxidant interactions with polyunsaturated fatty acids that promote lipid oxidation (Ramirez et al. 2005). On the other hand, there are studies in the literature showing that cooking does not affect lipid oxidation (Weber et al. 2008). This effect is explained by the reaction of the highly reactive compound measured by TBARS test with various compound such as proteins and amino acids present in the meat. Campo et al. (2006) mentioned a strong correlation between sensory perception of beef rancidity and TBARS value. For the oxidized taste–aroma of cooked beef meat, the threshold value of TBARS is reported to vary between 0.5 and 1 mg MDA kg−1 by experienced panelists, while it is reported to vary between 0.6 and 2 mg MDA kg−1 by inexperienced panelists (Juntachote et al. 2006; Cichoski et al. 2011). According to the results of the present study, it was observed that the use of basil in meatball production contributed to keeping the TBARS values of the samples close to the lower limit of 1 mg MDA kg−1.

Cooking loss values of the meatballs

Cooking loss values of the samples are also shown in Table 2. The use of basil at 1% in meatball production caused a significant decrease in the cooking loss value compared to the meatballs containing 0.25% and 0.5% basil and the control group meatballs. It is stated that the main cause of cooking loss as a result of the cooking process in meat and meat products is the water loss seen in the products during cooking (Rodriguez-Estrada et al. 1997). On the other hand, it is known that not only water but also some water-soluble substances are removed from the meat by cooking process (del Pulgar et al. 2012). Indeed, according to Gerber et al. (2009), Laroche showed that the meat juice moving away from the meat during cooking included a variety of compound such as myofibrillar or sarcoplasmic proteins, collagen, lipids, salt, polyphosphates and aroma compound. As the cooking temperature increased, the cooking loss values of the meatballs increased (p < 0.05). There are other studies in the literature showing that the cooking loss increases as the cooking temperature increases (Oz and Cakmak 2016; Tengilimoglu-Metin et al. 2017; Tengilimoglu-Metin and Kizil 2017).

Heterocyclic aromatic amine content of the meatballs

The recovery values of HAAs changed between 28.94 and 82.15%, while LOD (limit of detection = 3) and LOQ (limit of quantification = 10) values ranged between 0.004–0.025 ng g−1 and 0.013–0.085 ng g−1, respectively. In the present study, IQx, IQ, AαC and MeAαC compounds could not be detected, while the other HAAs were determined in varying amounts depending on the basil rate and the cooking temperature. The amounts of HAA in the meatballs are shown in Table 3.

Table 3.

The HAA content of the samples (ng g−1)

Cooking temperature (°C) Usage rate (%) IQx IQ MeIQx MeIQ 7,8-DiMeIQx 4,8-DiMeIQx PhIP AαC MeAαC Total HAA
150 0 ND ND ND ND ND ND ND ND ND ND
0.25 ND ND NQ ND ND ND ND ND ND NQ
0.5 ND ND NQ NQ ND ND ND ND ND NQ
0.75 ND ND NQ 0.06 ND ND ND ND ND 0.06
1 ND ND NQ 0.09 ND ND ND ND ND 0.09
200 0 ND ND 0.40 NQ ND ND ND ND ND 0.40
0.25 ND ND 0.41 NQ ND ND ND ND ND 0.41
0.5 ND ND 0.52 NQ ND ND ND ND ND 0.52
0.75 ND ND 0.92 NQ ND ND ND ND ND 0.92
1 ND ND 0.79 NQ ND ND ND ND ND 0.79
250 0 ND ND 0.63 0.09 0.08 0.08 0.58 ND ND 1.46
0.25 ND ND 0.62 0.07 NQ NQ ND ND ND 0.69
0.5 ND ND 1.52 0.09 NQ NQ ND ND ND 1.61
0.75 ND ND 0.93 0.08 NQ NQ ND ND ND 1.01
1 ND ND 0.53 0.07 ND ND ND ND ND 0.60
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ND Not detected, NQ not quantified

IQx could not be detected in the samples. Similarly, IQx was not detected by other researchers (Puangsombat and Smith 2010; Oz and Cakmak 2016). On the other hand, IQx was determined up to 0.45 ng g−1 by Oz et al. (2016) in beef chops cooked on a hot plate for 9 min at 250 °C and up to 3.48 ng g−1 by Tengilimoglu-Metin and Kizil (2017) in beef cooked in a pan at 150–250 °C for 10 min.

IQ could not be detected in the samples. Similarly, IQ was not detected by some researchers (Puangsombat and Smith 2010; Oz and Cakmak 2016; Oz et al. 2016). On the other hand, IQ was determined up to 0.60 ng g−1 by Ahn and Grun (2005) in beef cooked in a pan at 210 °C for 20 min and up to 11.29 ng g−1 by Lu et al. (2018) in deep-fat fried beef meatballs for 3 min at 180 °C.

MeIQx contents of the samples were determined up to 1.52 ng g−1. MeIQx was determined up to 6.51 ng g−1 by Ahn and Grun (2005) in beef cooked in a pan at 210 °C for 20 min and up to 7.80 ng g−1 by Cheng et al. (2007) in beef meatballs cooked in a pan at 200 °C for 12 min. On the other hand, MeIQx could not be determined by other researchers (Tengilimoglu-Metin et al. 2017; Lu et al. 2018; Nuray and Oz 2019).

MeIQ contents of the samples were determined up to 0.09 ng g−1. MeIQ was determined up to 1.15 ng g−1 by Ahn and Grun (2005) in beef cooked at 210 °C for 20 min, up to 6.35 ng g−1 by Tengilimoglu-Metin and Kizil (2017) in beef cooked in a pan for 10 min at 150–250 °C and up to 18.09 ng g−1 by Lu et al. (2018) in deep-fat fried beef meatballs for 3 min at 180 °C. On the other hand, MeIQ could not be detected by some researchers (Puangsombat and Smith 2010; Oz et al. 2016).

It is noteworthy that the use of basil in the production of meatballs leads to an increase in MeIQx and MeIQ content of some meatballs depending on the cooking temperature. It is known that Maillard reaction plays an important role in the formation of IQ-type HAAs (Jägerstad et al. 1983; Skog et al. 1998).

While MeIQx is formed by the reaction of dialkylpyrazine free radicals and creatine, MeIQ is formed by the reaction of alkylpyridine free radicals and creatine (Jägerstad et al. 1983). In addition, it is also stated that glycine and alanine play an important role in the formation of MeIQx (Skog et al. 1998) and MeIQ (Jägerstad et al. 1983), respectively. Glycine and alanine amino acids are also known to be the dominant amino acids of basil (Anonymous 2019). Therefore, it is thought that the basil used in the production of meatballs causes an increase in the content of MeIQx and MeIQ. In addition, it is thought that the basil used may have a pro-oxidant effect according to the usage rate or the antioxidant or prooxidant effect of basil may vary depending on the cooking temperature. On the other hand, it is also known that antioxidant substances can have prooxidant effects in vitro and in vivo systems depending on concentration, structure, test system and substrate (Nuray and Oz 2019).

7,8-DiMeIQx contents of the samples were determined up to – 0.08 ng g−1. The amount of 7,8-DiMeIQx in 1% basil added meatballs was below the LOD. 7,8-DiMeIQx was determined up to 0.15 ng g−1 by Oz and Cakmak (2016) in beef meatballs cooked on hot plate at 150 °C–250 °C for 9 min. On the other hand, 7,8-DiMeIQx could not be detected by some resarchers (Tengilimoglu-Metin et al. 2017).

4,8-DiMeIQx contents of the samples were determined up to – 0.08 ng g−1. While 4,8-DiMeIQx could not be detected in any of the samples cooked at 150 °C and 200 °C, the use of basil at the all use rates led to a decrease in the amount of 4,8-DiMeIQx in meatballs cooked at 250 °C. The amount of 4,8-DiMeIQx in 1% basil added meatballs was below the LOD. 4,8-DiMeIQx was determined up to 1.36 ng g−1 by Ahn and Grun (2005) in beef cooked in a pan at 210 °C for 20 min and up to 19.9 ng g−1 by Lu et al. (2018) in beef meatballs deep-fat fried for 3 min at 180 °C. On the other hand, 4,8-DiMeIQx could not be detected by some researchers (Tengilimoglu-Metin et al. 2017).

PhIP contents of the samples were determined up to – 0.58 ng g−1. While PhIP could not be detected in any of the samples cooked at 150 °C and 200 °C, the use of basil at the all use rates led to a decrease in the amount of PhIP in meatballs cooked at 250 °C. On the other hand, Damašius et al. (2011) found that usage of basil extract in beef cooked on grill at 200 °C for 20 min led to an increase in PhIP content compared to control group meat samples and the reason for this was explained by prooxidant effect of basil extract. PhIP was determined up to 2.24 ng g−1 by Oz et al. (2016) in beef chops cooked on hot plate at 200–250 °C for 9 min, up to 13.85 ng g−1 by Lu et al. (2018) in beef meatballs deep-fat fried for 3 min at 180 °C. However, there are also studies in which high levels of PhIP were determined in beef. Indeed, PhIP was determined up to 48 ng g−1 by Gross (1990) in samples cooked in a pan at 190 °C for 13 min. On the other hand, PhIP could not be detected by some researchers (Oz et al. 2016).

AαC could not be detected in the samples. Similarly, AαC was not detected by Tengilimoglu-Metin and Kizil (2017). On the other hand, AαC was detected up to 0.83 ng g−1 by Oz and Cakmak (2016) in beef meatballs cooked on hot plate at 200–250 °C for 9 min, up to 1.47 ng g−1 by Ahn and Grun (2005) in beef cooked in a pan at 210 °C for 20 min and up to 2.75 ng g−1 by Tengilimoglu-Metin and Kizil (2017) in beef cooked in a pan at 250 °C for 10 min.

MeAαC could not be detected in the samples. Similarly, MeAαC was not detected by some researchers (Viegas et al. 2012; Tengilimoglu-Metin and Kizil 2017). On the other hand, MeAαC was determined up to 0.05 ng g−1 by Oz and Cakmak (2016) in beef meatballs cooked on hot plate at 250 °C for 9 min and up to 0.48 ng g−1 by Tengilimoglu-Metin et al. (2017) in beef cooked in a pan for 10 min at 150–250 °C.

In the present research, it was determined that the total amount of HAAs in the meatballs samples analyzed was determined up to – 1.61 ng g−1 depending on the basil rate and the cooking temperature. While the total HAA contents of the meatballs cooked at 150 °C belonged to MeIQ, the total HAA contents of the meatballs cooked at 200 °C belonged to MeIQx. On the other hand, while the total HAA contents of the control group meatballs cooked at 250 °C consisted of MeIQx (43.15%), PhIP (39.73%), MeIQ (6.16%), 7,8-DiMeIQx (5.48%) and 4,8-DiMeIQx (5.48%), the total HAA contents of the meatballs containing different amounts of basil and cooked at the same temperature consisted of MeIQx (94.41–88.34%) and MeIQ (5.59–11.66%).

In terms of total HAA content, it is difficult to compare the results obtained in the present study with the data in the literature. There are two reasons for this. First; the present research is the first study to examine the effect of different amounts of basil in the production of meatballs on the formation of HAAs, secondly; it is known that cooking methods, temperature, duration, presence and amount of HAA precursors, meat type, etc. have an effect on HAA content (Oz and Kaya 2011a). Therefore, the differences are attributed to these factors. On the other hand, it is seen that the total amount of HAA calculated in meatballs analyzed in the present study is generally lower than in the literature.

Puangsombat and Smith (2010) investigated effect of usage of rosemary–water extract in different rates (0.05, 0.2 and 0.5%) in production of meatballs and different cooking temperatures (191 °C and 204 °C) on HAAs content of meatballs and found that usage of rosemary–water extracts decreased total amount of HAAs by 54.7–78.3%. Researchers reported that increase in cooking temperature increased total amount of HAA in all meatballs and total HAA content of meatballs ranged from 1.76 to 11.82 ng g−1.

Oz and Kaya (2011b) studied the inhibitory effect of red pepper on HAA content in beef (M. Longissimus dorsi) cooked at different temperatures (175 °C, 200 °C and 225 °C) and reported that usage of red pepper decreased total amount of HAA by 75.67 – 100%. Researchers reported that total amount of HAA in control group samples ranged from 2.63 to 9.47 ng g−1, while this amount was determined up to – 0.64 ng g−1 in the samples with red pepper. The authors declared that the reduction effect of red pepper on HAA formation could be due to the antioxidant activity of red pepper.

Zhu et al. (2016) evaluated inhibitory effects of dietary phenolic compound on HAA formation in beef meatballs with 8 different flavonoids and cooked at 230 °C for 20 min, it was reported that total HAA amount of control group meatballs was determined as 24.45 ng g−1, whereas in the flavonoid added meatballs, the total HAA amount ranged between 8.86 and 18.98 ng g−1.

Conclusion

In conclusion, the results obtained in the present study show that basil use in meatball production has both increasing and decreasing effect on both individual HAAs and total HAA content depending on the usage rate and cooking temperature. The inhibitory effect of basil on HAAs is thought to be due to the antioxidant effect of its phenolic compounds, while its increasing effect is attributed to the prooxidant effect. On the other hand, it is seen that even if 100 g of the meatballs containing 0.5% basil cooked at 250 °C whose total amount of HAA content is the highest, is eaten, the intake amount (0.161 μg) is far below the maximum acceptable daily consumption (15 μg day−1) reported by Oz (2019). The use of basil in meatball preparation has reduced the TBARS value, which affects the shelf life of meatballs. In light of the results obtained in the current study, it can be recommended that the use of 1% basil in the preparation of meatball that will be cooked at 250 °C. However, it is thought that studies examining the effects of basil use on sensory features that affect consumer acceptance are needed.

Acknowledgements

This research was supported by the Coordination Unit for Scientific Projects of Ataturk University with Project No: FYL-2018-6657. The financial support of Ataturk University is gratefully acknowledged.

Footnotes

Publisher's Note

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