Meat Science 104 (2015) 78–84

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A comparison of bleeding efficiency, microbiological quality andlipid oxidation in goats subjected to conscious halal slaughterand slaughter following minimal anesthesia

A.B. Sabow a,g, A.Q. Sazili a,f,⁎, I. Zulkifli a,b, Y.M. Goh b,c, M.Z.A. Ab Kadir e,h, N.R. Abdulla a,g, K. Nakyinsige f,i,U. Kaka d,k, K.D. Adeyemi a,j

a Department of Animal Science, Faculty of Agriculture, Faculty of Veterinary Medicine, 43400 UPM Serdang, Selangor, Malaysiab Institute of Tropical Agriculture, Faculty of Veterinary Medicine, 43400 UPM Serdang, Selangor, Malaysiac Department of Veterinary Preclinical Sciences, Faculty of Veterinary Medicine, 43400 UPM Serdang, Selangor, Malaysiad Department of Veterinary Clinical Studies, Faculty of Veterinary Medicine, 43400 UPM Serdang, Selangor, Malaysiae Department of Electrical and Electronic Engineering, Faculty of Engineering, 43400 UPM Serdang, Selangor, Malaysiaf Halal Products Research Institute Universiti Putra Malaysia, 43400 UPM Serdang, Selangor, Malaysiag Department of Animal Resource, University of Salahaddin, Erbil, Kurdistan Region, Iraqh Centre for Electromagnetic and Lighting Protection Research (CELP), Malaysiai Department of Food Science and Nutrition, Islamic University in Uganda, P.O. Box 2555, Mbale, Ugandaj Department of Animal Production, University of Ilorin, Ilorin, Nigeriak Department of Veterinary Surgery and Obstetrics, Faculty of Animal Husbandry and Veterinary Sciences, Sindh Agriculture University Tandojam, Sindh, Pakistan

⁎ Corresponding author at: Halal Products Research Ins43400 UPM Serdang, Selangor, Malaysia. Tel.: +60 3 894

E-mail addresses: awis@upm.edu.my, awisqurni@gma

http://dx.doi.org/10.1016/j.meatsci.2015.02.0040309-1740/© 2015 Elsevier Ltd. All rights reserved.

a b s t r a c t

a r t i c l e i n f o

Article history:Received 30 October 2014Received in revised form 10 February 2015Accepted 11 February 2015Available online 17 February 2015

Keywords:Halal slaughterMinimal anesthesiaBleeding efficiencyStorage stabilityGoats

The study assessed the effect of conscious halal slaughter and slaughter followingminimal anesthesia on bleedingefficiency of goats and keeping quality of goat meat. Ten Boer cross bucks were divided into two groups andsubjected to either halal slaughter without stunning (HS) or minimal anesthesia prior to slaughter (AS). Theblood lost during exsanguination was measured. Residual blood was further quantified by determination ofhemoglobin andmyoglobin content in longissimus lumborummuscle. Storage stability of themeatwas evaluatedby microbiological analysis and lipid oxidation. Blood loss at exsanguination, residual hemoglobin and lipidoxidationwere not significantly different (p N 0.05) betweenHS andAS. Lactic acid bacteriawas the onlymicrobethatwas significantly elevated after 24h of storage at 4 °C in theAS group. In conclusion, slaughtering goats underminimal anesthesia or fully conscious did not affect bleeding efficiency and keeping quality of goat meat.

© 2015 Elsevier Ltd. All rights reserved.

1. Introduction

Slaughtering animals for food is regulated by strict rules associatedwith safety and hygiene of food, working conditions and welfare ofanimals (Farouk et al., 2014). Slaughter practices have customarilymanaged components that affect wholesomeness and meat quality.Case in point, meat consumed by Muslims must be halal and thoyyib(acceptable and wholesome) (Nakyinsige et al., 2014). Themeat indus-try aims at attaining customer acceptability through the developmentand control of processes in order to produce wholesome productswith high quality and safety (Castro-Giráldez, Dols, Toldrá, & Fito,2011). Meanwhile, there exists consumer anticipation that meat prod-ucts should have the expected nutritional value, wholesomeness andfreshness, which are all affected by the animal production systems. In

titute Universiti Putra Malaysia,74870; fax: +60 3 89381024.il.com (A.Q. Sazili).

the production chain, slaughtering is a crucial step for animal welfare,meat quality and safety (Nakyinsige et al., 2014). Despite its short dura-tion, slaughter is an important critical point in the meat productionchain, with potential risks and its mishandling can ruin the effortsmade by producers during the longer growing and fattening phases(Anil, 2012). The amount of blood retained in meat is one of the mostcritical factors influencing the quality changes, contamination and dete-rioration (Ali, Abdalla, &Mahgoub, 2011). Blood is believed to be amag-nificent medium for bacterial growth. Blood components, particularlyhemoglobin, are compelling lipid oxidation promoters and may reducethe shelf-life of meat products (Alvarado, Richards, O'Keefe, & Wang,2007; Maqsood & Benjakul, 2011). Bleeding efficiency at sticking isinfluenced by: (1) blood vessels that are severed, (2) size and patencyof the sticking wound, (3) cardiac arrest at stunning, (4) orientation ofthe carcass-positioned horizontally or vertically, (5) vasodilation orvasoconstriction in the capillary bed, (6) tonic muscle contractionssqueezing blood capillaries and vessels, and (7) clonic activity causingmovement of blood toward the sticking wound (Gregory, 2005),

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which are all determined by the slaughter method (Nakyinsige et al.,2014).

Existing slaughter methods are comprehensively classified asconventional (procedures involving stunning) and religious (animalsare killed by neck cutting by use of a sharp knife in agreement withreligious prescriptions) or combination of conventional and religiouslike head-only electrical stunning followed by halal throat cut. Con-ventional slaughter methods focus around secular laws and entailstunning of animals before exsanguination. According to the EU CouncilDirective 93/119 (EU, 1993) and Council Regulation (EC) (No. 1099/2009) (EuropeanCommission, 2009) on animal protectionduring slaugh-ter, it is required that animals ought to be stunned with an intention tomake them unconscious before slaughter. Conversely, under traditionalreligious slaughter without stunning, animals must be executed by athroat cut so as to bring the animal to a quick death without agony,through severing of carotid arteries, jugular veins, trachea and esophaguspermitting rapid and complete bleeding (Anil, 2012; Anil & McKinstry,1991; Farouk, 2013). The halal method has been thought to provide con-siderable bleeding when the heart is still beating, whichmight be benefi-cial for shelf-life extension or meat quality maintenance (Addeen,Benjakul,Wattanachant, &Maqsood, 2014; Farouk et al., 2014). Althoughsome investigations have been conducted on the efficacy of differentslaughter methods on bleeding efficiency and meat keeping quality,most information originates from research in conventional slaughtermethods with limited comparison to specifically halal slaughter method.This was due to the limited access to religious slaughter without stunninginmost developed countries due to legal andwelfare reasons (Nakyinsigeet al., 2013). Animal subjected to minimal anesthesia has been identifiedas one of the bestmodel in terms of animalwelfare to study noxious stim-ulation associated with neck cut slaughter (Gibson et al., 2009; Gibson,Johnson, Stafford, Mitchinson, & Mellor, 2007), particularly in countrieswhere pre-slaughter desensitization and stunning are mandatory. Paincomprises both sensory and affective components (Fernandez & Turk,1992). Theminimally anesthetizedmethodwould have the added advan-tage of enabling the study of effects associated only with sensory painduring slaughter, versus both sensory and affective pain when animalsare slaughtered fully consciouswithout any formof stunning. It is possiblethat the presence of the affective pain componentswouldhave resulted inhigher cumulative pain-related stress in animals that are subjected toslaughter when they are fully conscious (i.e., halal slaughter). The inclu-sion of minimal anesthesia model would allow this experiment to studythe effect of physiological/sensory pain and its associated stress changeson muscle lipid oxidation, microbiological quality and exsanguination. Itis known that stress and pain typically have profound effects on the cen-tral nervous system and the mobilization of energy reserves. Thus, thisstudy aims to determinewhether slaughtering of minimally anesthetizedgoats (done to lessen slaughter pain) is comparable to non-anesthetizedhalal slaughter in terms of bleeding efficiency, oxidative stability and mi-crobiological quality.

2. Materials and methods

2.1. Ethical note

This study was conducted following the animal ethics guidelines ofthe Research Policy of Universiti Putra Malaysia.

2.2. Anesthesia and slaughter

A total of 10 male Boer cross goats weighing 23.15 ± 1.42 kg wereobtained from the same herd and of the same age (approx. 7 monthsold). The goats were allotted into two groups consisting of 5 animalseach and subjected to either conscious halal slaughterwithout stunningor pre-slaughter anesthesia followed by exsanguination. Slaughteringwas carried out at the Department of Animal Science research abattoir,Faculty of Agriculture, Universiti Putra Malaysia. In the halal method,

the animals were humanely slaughtered according to halal slaughteringprocedure as outlined in the MS1500: 2009 (Department of StandardsMalaysia, 2009). The process involved severing the carotid artery,jugular vein, trachea and esophagus. In the process of pre-slaughteranesthesia, animals were anesthetized using 5 mg/kg propofol adminis-tered by rapid injection into the cephalic vein and maintained with halo-thane in 100% oxygen, slaughtered and subsequently bled (Johnson et al.,2009; Kongara, Chambers, & Johnson, 2010).

2.3. Sample handling and storage

After evisceration, the longissimus lumborum (LL) muscle was sepa-rated into two parts, the first part was snap frozen in liquid nitrogen(Malaysian Oxygen Bhd., Malaysia) before being stored at −80 °Cuntil subsequent determination of TBARS at 0 day. The carcasses werehung in the cold room at 4 °C until the next samplingwas done at either1 or 7 days postmortem. On each sampling day, the chops were snapfrozen in liquid nitrogen and stored at −80 °C until subsequent deter-mination of TBARS at 1 and 7 d postmortem. The semitendinosusmusclewas aseptically divided into two portions. The first portionwas used formicrobial enumeration at d 0 while the other was packed in stomacherbags and stored at 4 °C for microbial enumeration at 1, 3 and 7 days.

2.4. Determination of blood loss

The amount of blood loss was estimated as the difference betweenpre-slaughter weight and post-slaughter weight (Velarde, Gispert,Diestre, & Manteca, 2003) once the animal is dead (based on ECG) asfollows:

Blood loss %ð Þ ¼ W1‐W2ð Þ �W1½ � � 100

Where:

W1 kgð Þ ¼ weight before slaughter

W2 kgð Þ ¼ weight after slaughter:

2.5. Quantification of heme proteins

2.5.1. Extraction of hemoglobin and myoglobinHemeproteins in the LLmuscleswere extracted following themeth-

od of O'Brien et al. (1992). For every sample, 5 g of pulverized muscletissue was mixed with 15 mL of ice cold extraction buffer containing80 mM KCl and 50 mM Tris–HCl at pH 8.0 and homogenized (WiggenHauser, Germany) for 40 s. Ice cold extraction buffer was used to pre-vent denaturation of myoglobin by acid produced during glycolysiswhereas KCl mimics intracellular ion concentration (O'Brien et al.,1992). The homogenates were rinsed with additional 5 mL of the bufferand centrifuged at 5000 g for 10min at 21 °C using a refrigerated centri-fuge machine (Avanti® J-26 XPI, Beckman Coulter®, USA) to clarify thesupernatant. The supernatants were aliquoted and stored at −80 °Cuntil total heme, hemoglobin and myoglobin evaluation.

2.5.2. Evaluation of hemoglobinHemoglobin was evaluated using a modified kinetic technique of

Goyal and Basak (2009) in which heme acts as a chemical catalyst tobreak down hydrogen peroxide intowater and nascent oxygen. Nascentoxygen then oxidizes o-tolidine to give an oxidized product a green-blue color. The rate of color development is directly proportional toheme concentration (Goyal & Basak, 2009). O-tolidine stock solutionwas prepared by dissolving 2 g of o-tolidine in 100 mL of solvent(20 mL of glacial acetic acid and 80 mL of ethanol) to create a stock so-lution. The working reagent (0.4 g/dL) was prepared by diluting stocksolution with the same solvent (1:5). A 100 μL of Triton-X-100 was

80 A.B. Sabow et al. / Meat Science 104 (2015) 78–84

mixedwith 100mLofworking reagent to increase the linearity of kinet-ic reaction. 100 milliliters of 2% (v/v) hydrogen peroxide solution wasprepared in deionized water and 2.26 g sodium acetate was added tocreate a buffering environment with glacial acetic acid present in thefinal reaction mixture so as to maintain pH between 3.0 and 3.5 in thefinal reaction mixture. The prepared solution was used within 6 to 8 h.

Hemoglobin standard solution was prepared by diluting bovineblood hemoglobin (H262G, Sigma- Aldrich, USA) in Tris–HCl buffer,pH 8.0, at a concentration of 240 mg/L. For the estimation of the enzy-matic reaction kinetics, the stock solution was diluted with 50 mMTris–HCl buffer to make final concentrations of 8.0, 12.0, 16.0, 20.0,24.0 28.0, 32.0, 36.0 and 40.0 mg/L of hemoglobin, respectively. Precise-ly, 1.0mL ofworking solution and 1.0mLof H2O2 solutionwere pipettedinto test tubes, mixed well and allowed to stand for 5 min at roomtemperature and 10 μL of each sample or standard was added to themixture. The absorbance was measured after 120 s (Cary 50 probeUV–visible spectrophotometer, Varian Australian, PTY LTD, Australia).

2.5.3. Myoglobin evaluationTwo milliliters of the supernatant were removed from the

−80 °C freezer, thawed, and 75% saturated with ammonium sulfate(0.525 g/mL) to precipitate hemoglobin while keeping myoglobin insolution. The solution was centrifuged at 2000 g at 21 °C for 45 minto separate the precipitated hemoglobin. The supernatant was filteredthrough a 0.45 μmdiameter filter (Denville® syringe filter). The myoglo-bin in filtrate was evaluated according to the method of Lerner (2009) inwhich heme acts as a chemical catalyst to break downhydrogen peroxideinto water and nascent oxygen (Goyal & Basak, 2009). Nascent oxygenthen oxidizes O-tolidine to give the oxidized product blue color. The sol-vent, O-tolidine stock solution, working reagent and hydrogen peroxidesolution were prepared as described in Section 2.5.2.

Myoglobin standard solution was prepared by diluting equine heartmyoglobin (Cat. No. M1882, Sigma-Aldrich, USA) with Tris–HCl buffer,pH 8.0, at a concentration of 250 mg/L. For the estimation of the enzy-matic reaction kinetics, the stock solution was further diluted with50 mM Tris–HCl buffer to make final concentrations of 10.0, 15.0, 20.0,25.0, 30.0 35.0 and 40.0mg/L ofmyoglobin, respectively. 200 μL ofmyo-globin filtrate or standard was mixed with 760 μL of deionized waterfollowed by 40 μL O-tolidine solution and 200 μL hydrogen peroxide so-lution. The mixture was then mixed and left for 2 min. Absorbance wasimmediately measured at 364 (Cary 50 probe UV–visible spectropho-tometer, Varian Australian, PTY LTD, Australia).

2.5.4. Total heme quantificationThe concentration of total heme proteinwas estimated in the super-

natants that were previously stored at −80 °C using the method ofO'Brien et al. (1992). The heme was oxidized with stock solution con-taining 100 μM K3Fe(CN)6. Ten microliters of stock solution was addedper mL of hemoprotein solution and its concentration evaluated bymeasuring absorbance at 540 and 580 nm and at the Soret band of420 nm (Cary 50 probe UV–visible spectrophotometer, VarianAustralian, PTY LTD, Australia). At these wavelengths oxidized equinemyoglobin had extinction coefficients of 3.19, 1.95 and 35.7 g−1 cm−1,respectively.

2.6. Lipid oxidation measurement

Lipid oxidation was measured as 2-thiobarbituric acid reactive sub-stances (TBARS) using QuantiChromTM TBARS Assay Kit (DTBA-100,BioAssay Systems, USA) following the manufacturer's description ofthe colorimetric protocol. Briefly, samples were manually pulverizedin liquid nitrogen. About 200 mg of the pulverized samples weremixed with 2 mL ice-cold phosphate buffered saline (PBS) and rapidlyhomogenized with an Ultra-Turrax T5FU (IKA-Labrortechnik Staufen,Germany) for 20 s on ice. Thereafter, 200 μL of homogenates weremixed with 200 μL of ice-cold 10% trichloroacetic acid (TCA) and

incubated on crushed ice for 5 min. This was followed by centrifugation(Eppendorf Centrifuge, Mikro 22R Hettich, Germany) at 14,000 g, 4 °Cfor 5 min. Standards were prepared by mixing 15 μL of the 1.5 mMmalondialdehyde (MDA) with 735 μL deionized water to obtain a finalconcentration of 30 μM MDA. Subsequently, 300, 180, 90 and 0 μL of30 μM MDA were diluted with 0, 120, 210 and 300 μL of deionizedwater to generate the final 30, 18, 9 and 0 μM MDA as standards 1, 2, 3and 4, respectively. Exactly 200 μL of samples and standards in labeled1.5 screw cap glass tubes, were added with 200 μL of thiobarbituricacid reagent and the mixture was incubated in a dry heating block(WiseTherm® HB, Germany) at 100 °C for 60 min. After equilibration toroom temperature, 100 μL of standards and samples was loaded in dupli-cate into wells of a clear flat- bottom 96-well plate (Greiner Bio-One,Germany). Finally, optical density (OD) was determined at 535 nm(OD535) using auto UV Xenon flash lamp microplate reader (infiniteM200, Tecan, Austria). After subtracting the OD of blank (standard4) from all standard and sample values, a standard curve was obtainedby plotting the ΔOD535 against standard concentrations.

TBARS (μM MDA equivalent) concentration of the samples wascalculated using the following equation:

TBARS ¼ R sample–R blankð Þ � Slope½ � � n

where,

Rsample and Rblank are the OD535nm of the sample and blank STD4ð Þ

n; the sample dilution factor n ¼ 3 for deproteinated samplesð Þ:

2.7. Microbiological analysis

On every sampling day (d 0, 1, 3 and 7), 5 g of meat samples fromsemitendinosusmuscle were aseptically weighed, transferred to a stom-acher bag containing 45 mL of 2.25% of peptone water (Merk KGaA,Germany) and homogenized using a stomacher (Inter Science, France)for 120 s at room temperature. For microbial enumeration, 100 μl sam-ples of 10-fold dilution in peptone water were spread on the surface ofdrymedia. Tenfold dilutionswere spread on petri dishes in duplicate forenumerations of total aerobic count (TAC) on Plate Count Agar (MerkKGaA, Germany), lactic acid bacteria on Man, Rogosa and Sharpe agar(Merk KGaA, Germany), Enterobacteriaceae on Violet Red Bile GlucoseAgar (Merk KGaA, Germany) and Pseudomonas spp. on CentrimideAgar (Merk KGaA, Germany). For all bacterial counts, plates were incu-bated at 32 °C for 72 h, except for Pseudomonas spp. which were incu-bated at 25 °C for 72 h (Bórnez, Linares, & Vergara, 2009). A colonycounter (Stuart®, USA) was used for counting.

2.8. Statistical analysis

The experiment followed completely randomized design. All analy-ses were performed using the GLM procedure of Statistical AnalysisSystem package (SAS) Version 9.2 software (Statistical Analysis System,SAS Institute Inc., Cary, NC, USA) and statistical significance was setat p b 0.05. Data were subjected to one-way analysis of variance(ANOVA) using a model that included the slaughter method as a possi-ble source of variation with sampling time as a repeated measure.Duncan's multiple range test was used to test the significance of vari-ance between the means of the studied parameters.

3. Results and discussion

3.1. Blood loss

Optimizing bleed out at slaughter and reducing carcass and meatdefects is a major goal of the meat processing industry since improved

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bleeding can improve the quality of the meat during storage (Ali,Lawson, Tauson, Jensen, & Chwalibog, 2007). Table 1 shows the resultof blood loss obtained from goats subjected to the halal slaughter (HS)and anesthesia with halothane (AS). There was no significant difference(p N 0.05) in blood loss between HS and AS. Though not statisticallydifferent, the rate of blood loss of HS was slightly higher than that ofAS. This could be because of the effect of increased heart rate immedi-ately after the neck cut in HS than AS group. Over the initial 90 s period,animals from the HS group showed a statistically (p b 0.05) higher rateof heartbeat than those from the AS groupwhich ranged from 86 to 187and 78 to 127 beats per minute for both treatments (HS and AS),respectively.

Studies on the effect of slaughtermethods on blood loss have yieldedcontrasting results. Our observation is in line with the finding ofAnil et al. (2004), Anil et al. (2006) and Agbeniga (2012) in which theslaughter method (conventional with stunning versus no stunning)did not affect blood loss in sheep and cattle. Anil et al. (2004) statedthat sheep slaughtered with or without head-only electrical stunninglost approximately 3.98% and 3.78% of their live-weight in blood at ex-sanguination respectively. In contrast, Velarde, Gispert, Diestre, andManteca (2003) assessed the effect of two methods of slaughteringused for halal meat production, no stunning and head-only electricalstunning on bleeding efficiency in lambs and found that the amount ofblood lost relative to body weight was significantly higher in stunnedcompared with non-stunned animals. They used light lambs (approxi-mately 19–21 kg live-weight), which produced a bleed out percentageof approximately 4.6% and 4.3% after neck cutting without stunning orelectrical stunning followed by neck cutting respectively. Warriss(1984) reported that sheep lost approximately 4% of their live-weightin blood at exsanguination. In the present experiment, the bleed outachieved by neck cutting of goats following minimal anesthesia orfully conscious was found to be higher (4.73–4.99%) than those report-ed above. This could be due to the orientation of the animal at slaughterwhich affects bleed-out or differences in techniques used for measure-ment of blood loss. Nakyinsige et al. (2014) found higher blood loss inrabbits subjected to halal slaughter without stunning as compared togas stunned animals followed by bleeding. Poor bleeding efficiency isa major quality defect which can even lead to undesirable discolorationand short shelf life (Griffiths & Purcell, 1984). Residual blood in carcassis essential in promoting microbiological deterioration of carcasses(Lerner, 2009; Warriss, 2000) and also an influential promoter of lipidoxidation as well as off-flavor (Nakyinsige et al., 2014).

3.2. Hemoglobin and myoglobin concentration

The residual hemoglobin and myoglobin concentrations in the LLmuscle obtained from goats subjected to different slaughter methodsare shown in Table 2. There was no significant difference (p N 0.05) inthe muscle residual hemoglobin concentration between the HS and ASgroup. This observation could be due to similarity in blood loss betweenthe two slaughter methods. The extent of vascular bed in the musclesand the bleeding of the carcass are limiting factors, which determinethe content of hemoglobin in meat (Oellingrath, Iversen, & Skrede,

Table 1Body weight pre-slaughter (LW), body weight post-bleeding and blood loss in goatssubjected to halal slaughter and pre-slaughter anesthesia followed by exsanguination(mean ± SE, n = 5).

Variable Treatment

HS AS

Body weight pre-slaughter LW (kg) 22.840 ± 1.663 23.450 ± 1.244Body weight post-bleeding (kg) 21.700 ± 0.457 22.342 ± 0.708Blood loss (kg) 1.140 ± 0.025 1.108 ± 0.040Blood loss (%) 4.991 ± 0.091 4.725 ± 0.071

HS: halal slaughter without stunning, AS: anesthesia with halothane followed byexsanguination.

1990). This observation is in line with the findings of Alvarado,Richards, O'Keefe, andWang (2007) who reported no significant differ-ence in the residual hemoglobin contents in the broiler breast musclebetween non-stunned and stunned broilers. Chrystall, Devine, andNewton (1981) reported that there was no difference in the residualblood content of lamb longissimus dorsimuscle using different slaughtermethods (Non-Stun Gash Cut (as traditionally practiced in NewZealand), Electric Head-Only Stun, Electric Head-to-Back Stun, Non-Stunned Neuromuscular Blocking Drug).

Themyoglobin concentration of the LLmuscles did not vary (p N 0.05)betweenHS and AS. Similarly, myoglobin content in broiler chickens sub-jected to different slaughter methods did not differ (Alvarado, Richards,O'Keefe, & Wang, 2007; Lerner, 2009). Also, the level of total heme(myoglobin+hemoglobin) in theHSmeatwas not significantly differentfrom the AS group. The concentration of heme protein content in meatdepends on the extent of vascular bed in themuscles aswell as the bleed-ing of the carcass (Nakyinsige, 2014;Oellingrath, Iversen, & Skrede, 1990).Thus, the total heme in goat LLmuscle can be a result of the loss of statis-tical significance in the residual blood in the muscles from HS and ASgroup. A similar trend was also reported by Chrystall, Devine, andNewton (1981)who indicated that therewas no effect of slaughtermeth-od on total pigment concentration (myoglobin and hemoglobin) inlambs' longissimus dorsimuscle.

3.3. Meat lipid oxidation

Lipid oxidation is a limiting factor which determines shelf life andsafety of meat (Insausti et al., 2001). It is considered a major cause ofnon-microbial meat spoilage, specifically under pro-oxidative con-ditions such as storage and cooking. It can also happen during refriger-ation and frozen storage (Soyer, Özalp, Dalmış, & Bilgin, 2010).The results of lipid oxidation levels during the first seven days postmor-tem are shown in Table 3. Slaughter method had no effect on goat meatlipid oxidation at 0, 1 and 7 d postmortem. These valueswere consistentwith the results for hemoglobin and myoglobin in longissimus lumborummuscle. Both hemoglobin and myoglobin are powerful promoters oflipid oxidation (Bekhit, Hopkins, Fahri, & Ponnampalam, 2013; Maqsood& Benjakul, 2011; Thiansilakul, Benjakul, Grunwald, & Richards, 2012).The dissociation of heme from myoglobin as well as iron fromheme is a major factor responsible for the pro-oxidative effect ofproteins and lipids (Faustman, Sun, Mancini, & Suman, 2010; Faustman,Yin, & Tatiyaborworntham, 2010). Conversely, hemoglobin is a strongerpro-oxidant than myoglobin and studies have shown that thiobarbituricacid-reactive substances (TBARS), peroxide values and hexanal duringstorage were greatest for hemoglobin as compared to myoglobin(Thiansilakul, Benjakul, Grunwald, & Richards, 2012). Moreover, residualblood in meat increases the concentration of heme proteins (mainlyhemoglobin) in meat. In general, lipid oxidation increased (p b 0.05)with aging time in both groups. However, no group (HS or AS) hadTBARS value that reached detectable concentration for humans asestablished by Insausti et al. (2001). The similarity in lipid oxidationobserved in the present study corroborates the report of Nakyinsigeet al. (2014) in rabbits,which showed that slaughtermethodhadnoeffect

Table 2Differences in hemoglobin, myoglobin and total heme of longissimus lumborum musclecontent in goats subjected to traditional halal slaughter without stunning and pre-slaughteranesthesia followed by exsanguination (mean ± SE, n = 5).

Variable Treatment

HS AS

Hemoglobin (mg/100 g) 0.851 ± 0.039 0.897 ± 0.051Myoglobin (mg/100 g) 337.196 ± 7.796 334.371 ± 11.104Total heme (mg/100 g) 339.049 ± 7.774 336.267 ± 11.067

HS: halal slaughter without stunning, AS: anesthesia with halothane followed byexsanguination.

Table 3Differences in the malondialdehyde content of longissimus lumborum muscle duringpostmortem aging periods in goats subjected to halal slaughter and pre-slaughteranesthesia followed by exsanguination (mean ± SE, n = 5).

Variable Postmortem agingperiods (day)

Treatment

HS AS

Lipid oxidation(mg MDA/kg meat)

0 0.498a,y ± 0.029 0.458a,y ± 0.0471 0.538a,y ± 0.018 0.561a,y ± 0.0297 1.139a,x ± 0.035 1.149a,x ± 0.055

HS: halal slaughter without stunning, AS: anesthesia with halothane followed byexsanguination.a,b means within the same row with different superscripts are significantly different(p b 0.05).x–z means within the same column with different superscripts are significantly different(p b 0.05).

a

aa

a

a

aa

a

0

0.5

1

1.5

2

2.5

3

3.5

4

4.5

5

0 1 3 7

Log

10 C

FU

/g

Days postmortem

Total aerobic count HS

AS

a

a

aa

a

a

aa

0

0.5

1

1.5

2

2.5

3

3.5

0 1 3 7

Log

10 C

FU

/g

Days postmortem

EntrobacteriaceaeHSAS

HS: Halal slaughter without stunning, AS: anesthesia withValues with different superscripts differ significantly at p<Values are means ± 1 standard error bar.

Fig. 1.Meatmicrobiological quality of cross Boer goats subjected to halal slaughterwithout stunstunning, AS: anesthesia with halothane followed by exsanguination. Values with different sup

82 A.B. Sabow et al. / Meat Science 104 (2015) 78–84

on meat lipid oxidation at 0 and 24 h postmortem. A similar trend wasalso reported by Linares, Berruga, Bórnez, and Vergara (2007) who indi-cated that no significant difference in lipid oxidation between electricallystunned and non-stunned lambs at 1 and 7 d postmortem.

3.4. Microbiological quality

Microbial contamination can reduce the quality of freshmeat, short-en its shelf life and result in economic loss and probably health hazards(Jouki & Khazaei, 2011). The microbiological quality of meat is influ-enced by the animal's physiological status at slaughter and the spreadof contamination during slaughter and processing (Koutsoumanis &Sofos, 2004). Fig. 1 shows microbial levels of chevon obtained fromgoats subjected to different slaughter methods. At d 0, microbial counts

a

aa

a

a

b a

a

0

0.5

1

1.5

2

2.5

0 1 3 7

Log

10 C

FU

/g

Days postmortem

Lactic acid bacteria HS

AS

aa

a

a

aa

a

a

0

0.5

1

1.5

2

2.5

0 1 3 7

Log

10 C

FU

/g

Days postmortem

Pseudomonas spp.HSAS

halothane followed by exsanguination.0.05.

ning and pre-slaughter anesthesia followed by exsanguination HS: Halal slaughterwithouterscripts differ significantly at p b 0.05. Values are means ± 1 standard error bar.

83A.B. Sabow et al. / Meat Science 104 (2015) 78–84

were not significantly different for the two slaughter methods. Howev-er, at d 1 postmortem, AS meat had higher (p b 0.05) growth of lacticacid bacteria while total aerobic counts, Enterobacteriaceae and Pseudo-monas spp. was unaffected by slaughter method. A faster pH declinecaused by minimal anesthesia causes aging to start earlier in the goatmeat as evident in our earlier study (Sabow et al., 2014), which wouldexplain the higher levels found for lactic acid bacterial growth in theAS group at 24 h. At d 3 and 7 postmortem, bacterial counts for all testedmicrobes were not affected by slaughter method. In general, increasedgrowth of all microorganisms with storage time was observed in meatsamples from both slaughter groups and meat samples from the pre-slaughter anesthesia animals had the highest counts of all microorgan-isms considered in this study. However, the levels were acceptable inall groups in all aging times as indicated by Insausti et al. (2001) andJeremiah (2001), that spoilage occurs when the levels of total viablecount and/or Enterobacteriaceae count reach 7–8 log cfu/g. Similarly,Nortjé and Shaw (1989) posited that spoilage occurs when the lacticacid bacteria count reaches 7 log cfu/g. In the present work, these levelswere not reached throughout the 7 d postmortem storage. The non-significant difference inmicrobiological quality between the treatmentsis expected given the similarity in the blood loss from the treatments.According to Nakyinsige et al. (2014), the amount of blood left withinthe carcass after bleeding is one of the most significant factors affectingthe level of contamination and thus increases the degree of the deterio-ration. The high nutritive value of blood in conjunction with favorabletemperature, pH, water activity and relative humidity influence thedegree of deterioration in meat (Lerner, 2009). For instance, glucosewhich is the substrate preferentially used by many microorganismssuch as Pseudomonas (Warriss, 2000) when growing inmeat is read-ily available in blood. Nakyinsige et al. (2014) observed that loweramount of residual blood in the carcass of rabbits subjected to halalslaughter caused lower bacteria count. In chicken, Ali, Abdalla, andMahgoub (2011) also reported that higher blood loss in halal slaugh-ter was associated with lower bacteria count in minced meat at 48 hpostmortem.

4. Conclusion

The result of the present study showed that blood loss or residualblood content and lipid oxidation and microbiological quality ofmeat from goats subjected to conscious halal slaughter is compara-ble to that from minimally anesthetized prior slaughter. However,lipid oxidation and the counts of all microorganisms of chevon increased(p b 0.05) with storage time in both groups. Thus, this study affirms thatslaughtering goats following minimal anesthesia did not result in poorbleed-out compared to slaughtering fully conscious goats and did notaffect the keeping quality of meat.

Conflict of interests

The authors declare that there is no conflict of interests regardingthe publication of this article.

Authors' contribution

ABS and GYM contributed to the idea, design, and execution ofthe study. ABS, KN and AQS performed the microbiological analysis,hemoglobin and myoglobin quantification, while KN, ZI, MAZAAQand AQS contributed toward the lipid oxidation measurement.KDA, UK and NRA assisted in all animal procedures for the experi-ment. GYM and ABS were responsible for the statistical analysis.All authors contributed equally to the write-up of the finalmanuscript.

Acknowledgments

The authors are very grateful to the Ministry of Education Malaysiafor the research fund provided through the Universiti Putra MalaysiaGrant (Project No. GP-IBT/2013/9409300).

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