The Potential Substitution of Oyster Shell Powder for Phosphate in Pork Patties Cured with Chinese Cabbage and Radish Powder

Chinese cabbage powder (CP) and RP, used as natural nitrate sources in this study, were prepared as previously reported by Jeong et al. (2020a). Vegetables, including Chinese cabbages and radishes, sourced from local markets in Korea, were processed into powders. The vegetables were cut into appropriate sizes and the inedible portions were removed. They were then sliced and washed before blending for 10 min using a food processor. The resulting mixture was packaged and frozen at −18°C. It was then dried for 12 h at 60°C, pulverized, and sieved using a 30-mesh. The dried vegetables were placed in vacuum-sealed packages and stored at −18°C until needed. Maltodextrin was added to the powders to standardize a nitrate content of 30,000 mg/kg and used on pork patties processing. OP was purchased from JK Biochem (Oyster Shell Calcium 40, Seoul, Korea), containing 52% calcium and less than 0.1% other minerals. The starter culture comprising Staphylococcus canosus was obtained from a supplier (Bactoferm^® S-B-61, Chr Hansen, Milwaukee, WI, USA). Sodium ascorbate (#35268) and sodium nitrite (S2252) were obtained from Sigma-Aldrich (St. Louis, MO, USA) and Acros Organics (Geel, Belgium), and acerola juice powder was procured from Diana Food (Antrain, France).

The preparation of patties necessitated the use of 15 kg of pork ham and back fat per batch of production. These materials were procured from a local supplier within 48 h of slaughter and their fat content was determined before processing. The amount of fat added was adjusted to attain the target fat content of 15% in the final product. Raw meat and back fat were subsequently processed using a chopper equipped with an 8 mm hole plate and a 4.5 mm hole plate, respectively, and then randomly assigned to six groups according to the formulation ratio (Table 1). Based on the weight of the ground pork and fat, basic ingredients, including 10% water, 1% NaCl, and 1% dextrose, were equally served in each batch and individually mixed using a mixer for 7 min. For the control (−), no additional additives were added beyond the basic ingredients, whereas the control (+), synthetic additives group, was supplemented with 0.3% STPP, 0.01% sodium nitrite, and 0.05% sodium ascorbate. In contrast, clean-label patties were tested using four different treatments. Treatment 1 included 0.4% CP and 0.3% OP, whereas treatment 2 contained 0.4% CP and 0.6% OP. Treatment 3 included 0.4% RP and 0.3% OP, whereas treatment 4 included 0.4% RP and 0.6% OP. The four treatments were supplemented with 0.04% starter culture to facilitate the conversion of nitrate to nitrite in the vegetable powders. Furthermore, these treatments received 0.295% acerola juice powder as a natural alternative to sodium ascorbate, serving as a curing accelerator. After mixing all batches, 90 g of each mixture was sequentially placed in Petri dishes (90×15 mm). All patties were then held at 40°C for 2 h for curing before freezing at −24°C. On the designated day of analysis, the patties were thawed at 5°C for 20 min prior to cooking and placed on an electric grill (GS6/C, Lincat, Lincoln, UK) preheated to 170°C for cooking per group. During cooking, the patties were flipped every two min they reached a core temperature of 75°C. After cooking, the samples were cooled to room temperature for one hour before analysis. This process was repeated thrice for both preparation and analysis.

The pH values were measured using a pH meter (Accumet^® AB150, Thermo Fisher Scientific, Singapore) following the method described by Yoon et al. (2023). The samples were blended with distilled water at a ratio of 1:5. Cooking loss was determined by measuring the weight of patties before and after cooking and expressing the difference as a percentage.

The shrinkage of patties was determined as described by Jeong et al. (2016). The initial thickness or diameter of the raw patties was measured using Vernier calipers (#530-101, Mitutoyo, Kawasaki, Japan). Following cooking and subsequent cooling, the patties were reassessed to determine the percentage reduction in their thickness or diameter relative to the original uncooked state.

The shear force of patties was evaluated according to the method described by Jeong et al. (2009). After cooking and cooling, the patties were divided into two sections measuring 2.5 cm in width. Five patties were used for each experimental group. The measurement obtained from the texture meter (TA-XT2i, Stable Micro System, Godalming, UK) equipped with a Warner-Bratzler blade, which indicates the maximum force required to shear the sample, was expressed in Newtons. The crosshead speed in the texture analyzer was determined to be 5 mm/s.

The patties were divided horizontally to their full height to evaluate the color of their cross-sections. The CIE L*a*b* values of the internal surfaces were measured using a color meter (CR-400, Konica Minolta, Osaka, Japan; illuminant C) equipped with an aperture (Φ 8 mm). Four readings were recorded for each treatment. A white standard plate (No. 20333081) was used to calibrate the color meter prior to the measurement.

The residual nitrite content in the cooked patties was analyzed using the AOAC method 973.31 (AOAC, 2016). A standard curve for sodium nitrite was employed to determine the nitrite content of the samples, and the results were reported in milligrams per kilogram.

The analysis of cured meat pigment and total pigments in cooked patties was conducted in accordance with the method outlined by Hornsey (1956), and the results were reported in milligrams per kilogram. The curing efficiency was calculated as the percentage of cured pigment in relation to the total pigment content, as described by King et al. (2023).

Thiobarbituric acid reactive substance (TBARS) analysis was conducted according to Tarladgis et al. (1960) to evaluate the malondialdehyde (MDA) content of the patties. The results obtained from TBARS analysis were presented in milligrams of MDA per kilogram of sample.

The present investigation was conducted using a completely randomized block design comprising six distinct groups. Data collected from three separate trials were statistically analyzed using the Proc GLM procedure within the SAS program (SAS Institute, Cary, NC, USA). A statistical model with a significance level of p<0.05 was applied to separate the mean differences between the groups for the dependent variables using Duncan’s multiple range test.

There were no differences (p>0.05) in the pH values between treatments 1 and 3 or between treatments 2 and 4 (Table 2). This finding indicates that the type of vegetable powder used did not have a notable influence on the pH of the pork patties. However, the treatments exhibited higher pH values (p<0.05) than those of the control samples (Table 2). The elevated pH observed in these patties may be due to the high pH (pH 9.84) of the OP added. Previous studies have suggested that the addition of oyster shell calcium to conventional meat products can cause an increase in pH (Cho and Jeong, 2018; Choi et al., 2014). This observation was corroborated by the present study using natural curing methods involving CP and RP. Additionally, as the amount of OP added to pork patties cured with vegetables increased, a significant increase (p<0.05) in the pH was observed (Table 2).

The use of OP in combination with CP or RP in cured pork patties resulted in lower (p<0.05) cooking loss than that of the control (−) (Table 2), suggesting that the addition of these natural additives contributed to increased water retention. In contrast, treatments 1–4, which used natural additives, demonstrated higher (p<0.05) cooking loss than the control (+) (Table 2). Previous research conducted by Choi et al. (2014) and Yoon et al. (2023) indicated that replacing phosphate with OP in pork ham or sausages did not result in a significant difference in cooking loss compared with conventional meat products. However, the findings from this study on pork patties differ from these previous results, suggesting that the replacement of phosphate with OP may have a different effect on cooking loss for this particular type of product. However, in this study, there was no difference (p>0.05) in cooking loss among treatments 2–4, but cooking loss was lower (p<0.05) than that in treatment 1 (Table 2). Therefore, the combination of 0.4% CP and 0.3% OP in clean-label patties was less effective at reducing cooking loss.

Phosphate is widely recognized for its capacity to minimize shrinkage of meat products during cooking (Anjaneyulu et al., 1990; Long et al., 2011). Interestingly, there was no significant difference (p>0.05) in the reduction of thickness and diameter between the control (+) with synthetic additives and all treatments with natural additives (Table 2). Moreover, the treatments showed a smaller reduction (p<0.05) in both diameter and thickness than the control (−) without any additives. These results suggest that OP may serve as an alternative to phosphate as it achieved a similar level of shrinkage as the control group (+) when incorporated into patties at a concentration of 0.3%, regardless of the vegetable powder used in combination.

Shear force refers to the force that a meat product can withstand when subjected to shearing, and serves as an indicator of its hardness (Novaković and Tomašević, 2017). Despite the variation in OP concentration and the type of vegetable powder used, no differences (p>0.05) were observed in shear force values between the different treatments and the control (+) (Table 2). However, among the various treatments, those containing 0.6% OP (treatments 2 and 4) had higher (p<0.05) shear force values than the control (−). In a previous investigation of pork sausages, Jeong (2018) found that a combination of 0.2% OP and 0.3% eggshell powder led to a significant improvement in hardness. Typically, the binding capacity of meat products is improved by increasing their ionic strength via the addition of phosphate and salt (Sebranek, 2009). In this study, the precise cause of the increase in shear force values owing to the use of OP is unknown. Nonetheless, it is conceivable that calcium in OP, functioning as a divalent cation, may facilitate protein bonding in meat, ultimately leading to the formation of a strong protein network (Cáceres et al., 2006; Lau et al., 2000). Although there was a trend towards higher shear forces in treatments 2 and 4 compared to treatments 1 and 3, statistical significance was not achieved (Table 2).

The control (+) showed no difference (p>0.05) in CIE L* values from all treatments or the control (−) (Table 3). However, the treatments showed lower (p<0.05) CIE L* values than the control (−). Additionally, the type of vegetable powder used and the amount of OP added did not have an impact (p>0.05) on the CIE L* values of pork patties within the treatment groups.

Efficient assessment of the curing process in meat products can be achieved by evaluating critical indicators, including CIE a* and cured pigment levels in cured meat (Feng et al., 2016; King et al., 2023). In this study, there was no significant difference (p>0.05) in the CIE a* values between the control (+) and all treatments (Table 3). Furthermore, the inclusion of OP in patties containing various vegetable powders did not affect (p>0.05) CIE a* values. Additionally, Lee et al. (2011) reported that adding OP to pork sausages did not result in a difference in CIE a* values compared with sausages containing phosphate. Although not statistically significant, the general pattern of CIE a* values decreased as the OP concentration in patties cured with CP or RP increased from 0.3% to 0.6% (Table 3). These findings may be attributed to the decrease in cured meat pigment, as shown in Table 4. As anticipated, the control (−) had the lowest (p<0.05) CIE a* values among all the patties tested because of the absence of nitrite or natural curing agents.

The control (−) exhibited the highest CIE b* values (p<0.05), whereas the control (+) had the lowest values (p<0.05; Table 3). However, among the treatments, vegetable powder type and OP did not significantly affect the CIE b* values (p>0.05). According to various studies, the use of plant-based powders as natural curing agents can affect the color of the final products, particularly when they originate from different sources (Jeong et al., 2020b; Riel et al., 2017; Sebranek and Bacus, 2007). In a related study, Jeong et al. (2020a) found that products using powders from leafy vegetables often exhibit higher levels of CIE b* compared to those using RP. Nevertheless, in this study, no significant difference (p>0.05) was observed in CIE b* values across the different vegetable sources used in the treatments (Table 3). One potential explanation for this result may be the incorporation of OP, which is typically white in color, along with CP or RP, resulting in a dilution effect on the CIE b* of the pork patties.

Treatments 1 to 4 had similar (p>0.05) residual nitrite content (Table 4). However, the treatments exhibited a lower (p<0.05) residual nitrite content than that of the control (+). This finding is consistent with several previous studies showing that clean-label or alternatively cured products generally have lower levels of residual nitrite than those containing sodium nitrite (Riel et al., 2017; Siekmann et al., 2021; Yong et al., 2021). This difference may be attributed to the reduction of nitrite to nitric oxide, which is influenced by the addition of a starter culture for alternative curing, resulting in its depletion (Ras et al., 2017; Wang et al., 2016). Another possible reason for the low levels of residual nitrite in meat products produced using alternative curing methods could be the presence of bioactive substances in plant sources (Viuda-Martos et al., 2010).

The results indicated that the amounts of cured meat pigment in treatments 2 and 4 were lower (p<0.05) than those in treatments 1 and 3 (Table 4). This outcome is attributed to the higher concentration of OP in pork patties, which significantly affects pH levels and thereby reduces cured meat pigment (Sebranek, 2009). Research has consistently emphasized the significance of pH in meat curing. Numerous studies, including those conducted by Fox (1966), Kim et al. (2019), and Sebranek (2009), have found that acidic conditions promote the reaction between reductants and nitrite, resulting in the rapid formation of nitric oxide, which is responsible for the coloration of cured meat. However, in this study, the relatively high pH in treatments 2 and 4, which contained 0.6% OP, delayed the nitrite curing reaction. Therefore, extending the curing time for these treatments may be necessary to produce sufficient amounts of nitric oxide (Sebranek, 2009). This may account for the lower CIE a* values observed in treatments 2 and 4 in our study (Table 3). However, treatments 1–3 did not differ (p>0.05) from the control (+) for the cured meat pigment (Table 4).

All four treatments showed significantly higher (p<0.05) total pigments than the control (+) (Table 4). The higher overall pigment level may be attributed to the influence of elevated pH levels in the treatments, which is in line with the findings of Fraqueza et al. (2006) and Hornsey (1959). Nevertheless, there were no significant differences (p>0.05) in total pigment content across all treatments or between the control groups (Table 4).

The results of this study indicate that the use of OP in curing meat patties has a negative impact on curing efficiency. Specifically, naturally cured patties containing varying levels of OP had a lower (p<0.05) curing efficiency than the control (+) (Table 4). Furthermore, among the treatments, treatments 2 and 4 had significantly lower (p<0.05) curing efficiency. This was likely due to the increased use of OP, as was observed for the cured meat pigment in this study. These results indicate that the effect of OP with high pH on the curing efficiency of pork patties is more pronounced than that of vegetable powders in the study conducted. According to Sebranek (2009), elevated pH levels can negatively affect the curing process of meat by slowing down the reaction.

As expected, the highest TBARS values (p<0.05) were observed in the control (−) (Table 4). However, the combination of CP, RP, and OP in pork patties did not have a significant impact (p>0.05) on TBARS values compared to the control (+). This is consistent with the findings of Yoon et al. (2023), who reported similar TBARS values for pork sausages cured with 100 ppm sodium nitrite or 0.4% RP, and suggested that the antioxidative activity of nitrite converted from the added nitrate source, RP, may have inhibited lipid oxidation. Furthermore, Yoon (2021) investigated the efficacy of citrus fiber, dried plum powder, and oyster shell calcium as alternatives to phosphates in pork sausages. The results indicate that oyster shell calcium exhibited no significant differences in TBARS values throughout the storage period, suggesting an antioxidant effect attributable to the presence of CaO. Therefore, our results suggest that the natural ingredients used in clean-label pork patties effectively inhibit lipid oxidation in the final product.