Effect of Teucrium polium Essential Oil on the Physicochemical and...

EFFECT OF TEUCRIUM POLIUM ESSENTIAL OIL ON THEPHYSICOCHEMICAL AND SENSORY PROPERTIES OFPROBIOTIC YOGHURTRAZZAGH MAHMOUDI1,3, PAYMAN ZARE2, PARVIZ HASSANZADEH1and SOMA NOSRATPOUR1

1Department of Food Hygiene and Aquatics, Faculty of Veterinary Medicine, University of Tabriz, Tabriz PO Box 5166614779, Iran2Department of Pathobiology, Faculty of Veterinary Medicine, University of Tabriz, Tabriz, Iran

3Corresponding author.TEL: +98 (0) 411 3392378;FAX: +98 (0) 411 3357834;EMAIL: [email protected]

Received for Publication July 15, 2012Accepted for Publication September 30, 2012

doi:10.1111/jfpp.12042

ABSTRACT

Yoghurt samples treated with Teucrium polium essential oil (EO; 40, 60 and80 ppm) and Lactobacillus casei (108–109 cfu/mL) were stored up to 28 days at 4C.The physicochemical, viability of L. casei and organoleptic properties of yoghurtsamples were analyzed at a 7-day interval during storage. A gradual increase inacidity was observed, whereas a decline in pH and solid nonfat contents was notedin all the samples (P < 0.05). Changes in protein and fat content of yoghurtsamples during storage period were not remarkable. The survival of L. caseidecreased throughout the storage period. Nevertheless, probiotic yoghurt andtreatment containing 60 ppm EO had the highest (P < 0.05) viable count of probi-otic bacteria (6.44cfu/g). Considering the organoleptic effect of the EO, the besttreatment with acceptable flavor, good appearance without any signs of spoilageorganisms and also with appropriated viability of L. casei was obtained at combi-nation of 40 ppm EO and probiotic.

PRACTICAL APPLICATIONS

Based on our results, yoghurt can be a very suitable food product to carry relevantprobiotic bacteria while adding certain herbal EOs. In this study, T. polium EOhad adverse effect neither on physicochemical properties nor on the viability ofL. casei of yoghurt. In addition, T. polium EO is nontoxic and safe for publichealth, so it can be an acceptable alternative to chemical preservatives.

INTRODUCTION

Milk and dairy products are consumed all over the world.Of these, yoghurt is the most popular and unique, which isbeing enjoyed for its refreshing taste and beneficial proper-ties. It possesses high nutritional, antagonistic and thera-peutic values; yoghurt provides higher levels of protein,carbohydrate, calcium and certain B vitamins than milk(Abd-EI Fattah et al. 2010). The properties of yoghurt aredue to the presence of lactic acid bacteria (LAB) in it, whichferment lactose to lactic acid that improves the nutritionaland therapeutic values of yoghurt (Bylund 1995; Adolfssonet al. 2004). Probiotic bacteria defined as living microorgan-isms, which upon ingestion in certain numbers, exert healthbenefits beyond inherent basic nutrition, have become amajor topic of LAB research over the past 20 years (Guarnerand Shaafsma 1998). Probiotics have been defined as live

microbial food supplements that benefit human health(McFarland 2000; Salminen 2001). More than 90% of pro-biotic products contain various species of Lactobacilli andbifidobacteria (Shah 2001). Viable LAB of probiotic foodshave several scientifically established and/or clinicallyproved health effects, such as reduction and prevention ofdiarrheas of different origin, improvement of the intestinalmicrobial balance by antimicrobial activity, alleviation oflactose intolerance symptoms, prevention of food allergy,enhancement of immune potency and antitumorigenicactivities (McFarland 2000; Andersson et al. 2001; Salminen2001). Moreover, some studies have shown that certainLAB possess antioxidative activity (Kullissar et al. 2002;Songisepp et al. 2004).

Different types of products were proposed as carrierfoods for probiotic microorganisms by which consumerscan take in large amounts of probiotic cells for the thera-

Journal of Food Processing and Preservation ISSN 1745-4549

1Journal of Food Processing and Preservation •• (2012) ••–•• © 2012 Wiley Periodicals, Inc.

peutic effect. Yoghurt as one of them products consumptionhas increased significantly, presumably because of its per-ceived health benefits (Wood 1992). In recent past, the useof different fruit flavors in yoghurt manufacturing has beenattempted increasingly. Several workers reported the chemi-cal composition and microbial quality of different fruit-flavored yoghurts. Con et al. (1996) found that the additionof fruit flavor had no significant effect on the total bacterialand coliform counts. A few studies have also been reportedon the effect of alternative sweeteners on the yoghurtquality. Mc Gregor and White (1986) reported that high-fructose corn syrup has increased the acceptability ofyoghurt. One of the most accepted ways to extend the shelflife of perishable food products is through the use of bio-preservatives (Burt 2004; Draughon 2004). Essential oils(EOs) are aromatic, oily liquids obtained from plant materi-als. Steam distillation is the most commonly used methodfor commercial production of EO. It has long been recog-nized that some EOs have antimicrobial properties (Burt2004) and that they can be used as food flavoring agents orpreservatives, and for medicinal purposes (El-Nawawy et al.1998).

Teucrium polium L. locally named Kalpooreh has beenknown as an important traditional medicinal plant inKhuzestan, South West of Iran. T. polium L. is the memberof Lamiaceae family, a grass plant, durable, with 10–30 cmin height and callous with exterior that ordinarily have dis-persal in rocky and sandy area of Europe zones, North ofAfrica and Southwest of Asia like Iran. Medical of reputa-tion of this plant was noticed in traditional medicine bySocrates and Jalinous (Zargari 1994). Researchers showedthat this plant have antidiabetic, anti-inflammatory, antis-pasmodic, analgesic and antioxidant effects (Shahraki et al.2006; Hasani et al. 2007).

This study aimed to evaluate the chemical components ofT. polium L. EO, effect of this EO at different concentrationson the physicochemical, organoleptic properties of the pro-biotic yoghurt and the viability of the Lactobacillus casei wasalso investigated.

MATERIAL AND METHODS

Extraction and GC/MS1 Analysis ofT. polium EO

Plant Material. The T. polium L. was collected duringflowering stage from West of Iran (Kerman province) andidentified by the herbarium of Faculty of Pharmacy, Univer-sity of Tabriz, Tabriz, Iran, then shade dried and grindedinto powder. The prepared powder was kept in tight con-tainers protected completely from light.

Preparation of the EO. Dry aerial parts (100 g) ofT. polium L. were subjected to the hydrodistillation of2.5 h, using a clevenger-type apparatus, according to themethod recommended by the European Pharmacopoeia(Maissoneuve 1983) to produce oils. Light yellow-coloredoil was obtained, which was dried over anhydrous sodiumsulfate and stored at +4C until tested and analyzed.

GC-MS Analysis of the EO. The EO was analyzed byGC. The chromatograph (Agilent 6890; Agilent Technolo-gies, Kansas) was equipped with an HP-5MS capillarycolumn (30 ¥ 0.25 mm ID ¥0.25 mm film thickness) andthe data were taken under the following conditions: initialtemperature 50C, temperature ramp 5C/min, 240C/min to300C (holding for 3 min) and injector temperature at 290C.The carrier gas was helium and the split ratio was 0.8/mL/min. For confirmation of analysis results, EO was also ana-lyzed by GC-MS (Agilent 6890 gas chromatographequipped with an Agilent 5973 mass-selective detector;Agilent, U.K.) and the same capillary column and analyticalconditions as above. The MS was run in electron ionizationmode with ionization energy of 70 eV.

Starter and Probiotic Bacteria. Freeze-dried yoghurtinoculants (R 704, Christian Hansen Co., Horsholm,Denmark) containing Streptococcus salivarius ssp. thermo-philus and L.delbrueckii ssp. bulgaricus (1:1) was used as astarter. A commercial lyophilized culture of the probioticL. casei ATCC 3939 was obtained from the Iranian Organi-zation of Industrial Research. Subcultivation and prepara-tion of the probiotic bacteria were conducted according toPhillip et al. (2006).

Preparation and Inoculation of Yoghurt. Raw cowmilk (pH: 6.67, protein: 3.04, fat: 1.85, solid nonfat [SNF]:8.4, total solid [TS]: 9.62) was subjected to a heat treatmentat 90C for 20 min, followed by cooling to 40–45C. The EOwas added to milk before processing with different concen-trations (40, 60, 80 ppm) followed by mixing. As starterculture, yoghurt (L. bulgaricus and S. thermophilus) wasadded (1.5%) to the milk, followed by mixing, finallyL. casei (108–109 cfu/mL) was added, then packed in steril-ized glass-capped cups 250 mL capacity, followed by incu-bation at 40C for 3 h until gel forms (pH 4.5). Freshyoghurt was cooled and stored at refrigeration at 4C untilexamination to slow down the physical, chemical and sur-vivability of L. casei analysis. Yoghurt samples include:sample A: control, sample B: probiotic yoghurt, sample C:probiotic yoghurt with EO80 ppm, sample D: probioticyoghurt with EO 60 ppm, sample E: probiotic yoghurt withEO 40 ppm, sample F: yoghurt with EO 80 ppm, sample G:yoghurt with EO 60 ppm and sample H: yoghurt EO40 ppm.1 Gas chromatography mass spectrophotometer.

USE OF THE ESSENTIAL OIL IN PROBIOTIC YOGHURT R. MAHMOUDI ET AL.

2 Journal of Food Processing and Preservation •• (2012) ••–•• © 2012 Wiley Periodicals, Inc.

Physiochemical Analyses

pH and Titratable Acidity. The pH values of yoghurtsamples were measured using a digital pH meter (Nick, 776,Jena, Germany) fitted with a standard, combined glass elec-trode. The pH meter was calibrated with buffer solutions ofpH 4 and pH 9 prior to use, The titratable acidity was deter-mined by the method reported by Iwalokun and Shittu(2007), which is in accordance with IDF (1991) standards.Five grams of yoghurt sample was titrated with 0.1 N NaOHsolutions using 1% phenolphthalein as indicator. Thetitratable acidity was calculated as grams of lactic acid in100 g of yoghurt using the following equation: titratableacidity = (V ¥ 0.9)/m.

Where V is the volume (in mL) of 0.1 N NaOH solutionconsumed, m is the mass (in grams) of test portion ofyoghurt sample and 0.9 is the correction factor for lacticacid.

Protein, Fat and SNF content. The protein contentswere determined by the micro Kjeldal method according toAOAC (2000). The fat content was determined by the modi-fied Mojonnier ether extraction method (AOAC 1995). Theextracted fat is dried to a constant weight and expressed aspercent fat per weight.

SNF was determined by conducting total solids (AOAC1995) and fat analyses. Percent fat was subtracted frompercent total solids to obtain percent solids-not-fat.

Assessing Viability of L. casei. Survival of probioticbacteria during storage period of yoghurt samples (ondays 1, 7, 14, 21 and 28) was conducted according tostandard method. One gram of yoghurt sample werehomogenized aseptically in a stomacher with 9 mL ofsterile peptone water (Merck, Darmstadt, Germany)(0.1%) and 10-fold (102–108) serial dilutions were pre-pared. The enumeration was carried out using spreadplates with a 100 ppm inoculum on reinforced clostridialagar (RCA) with bromocresol green and vancomycinmedium (RCABV). The pH of the RCA agar base wasadjusted to 5.5 prior to autoclaving and then bromocresolgreen stock 0.2% w/v (prepared as previously described)added at the rate of 20 mL/L. Vancomycin stock solution(2% w/v) was prepared with distilled water and filter-sterilized through a 0.45-mm membrane. This was added at

the rate of 0.5 mL/L to the molten agar. The plates wereincubated anaerobically in gas jars using the GasPakSystem, (Thermo Fisher Scientific Inc., Watham, MA) for48 h at 37C prior to observation. All plate counts werecarried out in triplicates. Plates containing 25–250 colonieswere enumerated and recorded as colony forming units(cfu/g) of the product (Phillip et al. 2006).

Sensory Analysis

The sensory effects of adding of T. polium L. EO and probi-otic bacteria to yoghurt were evaluated using an acceptancetest. Yoghurt samples with various amounts of EO asdescribed previously were equally divided into seven partsof 20 g each and placed on white plates coded with three-digit random numbers. The sensory evaluation was per-formed by a panel of seven judges consisting of thescientific staff of the Department of Food Hygiene, Facultyof Veterinary Medicine, University of Tabriz, experienced inthe sensory analysis of food. Each panelist evaluated thesamples by rating them using a 9-point scale (Table 1),where 9 = like extremely and 1 = dislike extremely, forvarious characteristics such as (appearance) color, odor andflavor (Meilgaard et al. 1999).

The variability of acceptance or liking of the samples wasanalyzed by analysis of variance (ANOVA) and Fisher’s leastsignificant difference procedure, using the SPSS 17 statisti-cal software package (SPSS 17 for Windows; SPSS, Inc.,Chicago, IL).

Statistical Analyses

All statistical analyses were performed using software 17SPSS. Data related to the mean values of microbial countsand physicochemical evaluations were subjected to ANOVA.Significant results were considered at P < 0.05. All experi-ments were performed in triplicate.

RESULT AND DISCUSSION

Chemical Analysis of the T. polium EO

The EO was extracted by the hydrodistillation of the driedaerial parts of T. polium and yield of the EO was 0.5%. GCand GC-MS analysis enabled the identification of a total of21 constituents, representing 90.40% of the oil. The relative

TABLE 1. ACCEPTANCE TEST (9-POINT SCALE)

Rating ofscale 1 2 3 4 5 6 7 8 9

Properties Dislikeextremely

Dislikevery much

Dislikemoderately

Dislikeslightly

Neither likenor dislike

Likeslightly

Likemoderately

Likevery much

Likeextremely

R. MAHMOUDI ET AL. USE OF THE ESSENTIAL OIL IN PROBIOTIC YOGHURT


concentrations of the volatile components identified arepresented in Table 2, according to their retention time andthe relative concentrations of the identified volatile compo-nents (%). The main components found as spathulenol(15.06%), beta-pinene (11.02), beta-myrcene (10.05), ger-macrene B (10.11%), germacrene D (8.15%), bicyclogerma-crene (8.25%) and linalool (4.02). Predominantly, the oilwas rich in sesquiterpenoids.

The EO of Teucrium species has been studied in Iranand in the world. A scientific study on the T. stocksianumsubsp. Stocksianum EO in April 2003 found 38 compoundswith 97.7% of the oil were identified. The major componentof the oil as camphene (20.6%), alpha-pinene (19.7%),myrcene (10.2%) and carvacrol (9.90%; Jaimand et al.2006). The studies made have reported the volatile constitu-ents of T. flavum leaves from Iran at the full flowering stagein May 2001, 18 compounds were identified constitutingabout 99% with 0.2% yield of the oil that major compo-nents were b-caryophyllene (30.7%), germacrene (21.3%)and alpha-humulene (8.4%) (Bahar and Mirza 2003).

A study on the EO obtained from T. polium grow inIran (Eikani et al. 1999) revealed the presence of sesquiter-penes as major components, germacrene D (13.2%),b-caryophyllene (18%), spathulenol (10.4%) and bicy-clogermacrene (9.0%) being the major componentsidentified.

Thirty-seven components were detected in the EOobtained from the aerial part of Turkish T. polium (Cakiret al. 1998). The major components identified were

beta-pinene (18%), b-caryophyllene (17.8%), alpha-pinene(12%), caryophyllene oxide (10%), myrcene (6.8%), germa-crene D (5.3%), limonene (3.5%) and spathulenol (3.3%).It is the first time that a T. polium has been found to containspathulenol (14.65%) at such high percentage; spathulenolwas reported as an important constituents of T. polium(10.4%; Eikani et al. 1999) and in the oil obtained from theaerial parts of Turkish T. polium (3.3%; Cakir et al. 1998).Based on the results of GC/MS analysis of T. polium EO inour study, beta-pinene, beta-myrcene, germacrene B, ger-macrene D, bicyclogermacrene and linalool were the majorcomponents and also these component has been reportedas the major compounds of EO obtained from Teucriumspecies in other scientific researches (Cakir et al. 1998;Eikani et al. 1999; Bahar and Mirza 2003; Aburjai et al. 2006;Amiri 2008).

It is possible to say that the difference in the quality orquantity of the composition of volatile oils may be due togenetic, differing chemotype, drying conditions, mode ofdistillation and or extraction and geographic or climaticfactors.

Physicochemical Properties

The results showing the effect of storage and treatments onthe pH values of various yoghurt samples are presented inTable 3. A gradual and consistent decrease in pH along thestorage was noted (especially from the first day until the14th day); also there was no significant change in pH 14th

TABLE 2. COMPOSITION OF T. POLIUM EOAS DETERMINED BY GC-MS

Compound no. Compound name RT (min) Percentage (%)

1 Verbenene 6.97 1.252 Beta-pinene 7.59 11.023 Beta-myrcene 7.93 10.054 Benzene, 1-methyl 8.88 1.665 Cyclohexene, 1-methyl-4-(1-methylethenyl) 9.01 2.006 2,3,3-Trimethyl-3-cyclopentene acetaldehyde 11.80 1.717 Bicyclogermacrene 12.49 8.258 Benzene 13.87 1.469 Bicyclo[3.1.1]hept-3-en-2-one, 4,6,6-trimethyl 14.27 1.31

10 1,6,10-Dodecatriene, 7,11-dimethyl-3-methylene 20.71 1.2311 Germacrene D 21.42 8.1512 Germacrene B 23.28 10.1113 sesquisabinene hydrate 23.46 5.2614 Spathulenol 24.15 15.0615 o-Menth-8-ene 24.63 1.2316 3-Cyclohexene-1-methanol, .alpha.,4-dimethyl 24.75 1.2317 1H-3a,7-Methanoazulene 25.12 1.8418 Naphthalene 25.81 1.2119 Linalool 25.97 4.0220 Cyclolongifolene oxide, dehydro 26.22 1.0521 Benzenemethanol, 4-(1-methylethyl) 26.98 1.30

Total 90.40

RT, retention time (min).



day to end storage period. The initial pH values for the dif-ferent yoghurt types ranged from 4.45 to 4.60. In addition,the different concentration of T. polium EO did not seem toaffect the pH of the samples significantly. The above resultsare in line with the findings of Singh et al. (2011), Yegane-hzad et al. (2007) and Wolfschoon (1983); they reported adecrease in pH of yoghurt during storage. The observed pHrange of 4.60–3.79 in our yoghurt samples is still within therange reported by earlier workers (Salvador and Fiszman2004; Haddadin 2005; Muhammed et al. 2005).

The change in total acidity (TA) is a very importantfactor since it affects the shelf life and the acceptability ofyoghurt (Al Otaibi and El Demerdash 2008). The titratableacidity values of different curd samples (Table 4) increasedduring the cold storage of yoghurt. However, during theinitial days of storage, additives have not significantlyaffected the titratable acidity, but a significant (P < 0.05)

difference in titratable acidity of control and treated sets(especially combination EO and probiotic treatments) wasobserved on later days. The highest values were obtainedwith yoghurt samples containing 60 ppm of the EO andL. casei when fresh and it increased up to the end of storage(day 28), suggesting that the EOs had a stimulatory effect onthe starter culture and L. casei.

These results were in agreement with that obtained byAl-adamany et al. (2003), Shin et al. (1991), Abbas andOsman (1998), Al Otaibi and El Demerdash (2008), Singhet al. (2011), they reported that the TA increased graduallyduring storage period.

The reason for the decrease in pH is a function of aciditythat was increased during the storage period because ofthe conversion of lactose to lactic acid. Taken together, itappears that the composition of starter culture, fermenta-tion temperature, storage duration, contamination, etc.,

TABLE 3. CHANGES IN pH VALUE OFYOGHURT SAMPLES DURING THE STORAGE Yoghurt

samples

Storage period(days)

1 7 14 21 28

pHA 4.60 � 0.03a 4.32 � 0.07b 3.95 � 0.09c 3.95 � 0.03c 3.90 � 0.02c

B 4.51 � 0.01a 4.35 � 0.09b 3.79 � 0.03d 3.77 � 0.06d 3.71 � 0.04d

C 4.47 � 0.01a 4.10 � 0.12c 3.80 � 0.05d 3.75 � 0.09d 3.72 � 0.07d

D 4.46 � 0.02a 3.99 � 0.07c 3.95 � 0.07c 3.84 � 0.13d 3.80 � 0.05d

E 4.50 � 0.05a 4.00 � 0.15c 3.90 � 0.10c 3.81 � 0.18d 3.80 � 0.10d

F 4.52 � 0.09a 4.22 � 0.02b 3.96 � 0.14c 3.89 � 0.05d 3.81 � 0.18d

G 4.48 � 0.07a 4.06 � 0.11c 3.86 � 0.06d 3.84 � 0.03d 3.80 � 0.04d

H 4.45 � 0.04a 4.06 � 0.08c 4.00 � 0.02c 3.92 � 0.07c 3.85 � 0.07d

The mean values followed by the same letter in the column are nonsignificantly different(P < 0.05).Sample A, yoghurt with no additive; sample B, probiotic yoghurt; sample C, probiotic yoghurtwith 80 ppm EO; sample D, probiotic yoghurt with 60 ppm EO; sample E, probiotic yoghurt with40 ppm EO; sample F, yoghurt with 80 ppm EO; sample G, y with 60 ppm EO; sample H, yoghurtwith 40 ppm EO.

TABLE 4. CHANGES IN TA% OF THEYOGHURT SAMPLES DURING THE STORAGE Yoghurt

samples


1 7 14 21 28

Titration acidityA 0.84 � 0.03a 0.88 � 0.05a 0.95 � 0.08b 0.95 � 0.10b 0.97 � 0.07b

B 0.82 � 0.09a 0.85 � 0.04a 0.94 � 0.15b 1.04 � 0.05b 1.08 � 0.10b

C 0.80 � 0.05a 0.88 � 0.12a 1.08 � 0.09b 1.16 � 0.11c 1.160 � 0.08c

D 0.80 � 0.13a 0.87 � 0.09a 1.06 � 0.06b 1.14 � 0.08c 1.190 � 0.14c

E 0.78 � 0.10a 0.85 � 0.04a 1.00 � 0.11b 1.08 � 0.06b 1.100 � 0.05c

F 0.79 � 0.11a 0.84 � 0.18a 0.95 � 0.09b 0.96 � 0.10b 0.98 � 0.13b

G 0.77 � 0.07a 0.84 � 0.10a 1.03 � 0.06b 1.05 � 0.11b 1.09 � 0.07b

H 0.75 � 0.08a 0.80 � 0.05a 0.93 � 0.08b 0.96 � 0.06b 0.99 � 0.09b

The mean values followed by the same letter in the column are nonsignificantly different(P < 0.05).Sample A, yoghurt with no additive; sample B, probiotic yoghurt; sample C, probiotic yoghurtwith 80 ppm EO; sample D, probiotic yoghurt with 60 ppm EO; sample E, probiotic yoghurt with40 ppm EO; sample F, yoghurt with 80 ppm EO; sample G, yoghurt with 60 ppm EO; sample H,yoghurt with 40 ppm EO.



could influence the overall level of acidity and pH of storedyoghurt samples (Singh et al. 2011).

The effect of storage and treatments on the SNF contentsof various yoghurt samples is shown in Table 5. A gradualreduction in SNF throughout the storage period wasobserved, which might be due to the conversion of lactoseto lactic acid by lactose-fermenting bacteria (Anjum andZahoor 2007). Lactose forms a considerable amount ofsolids in milk. Lactose fermentation into lactic acidcan reduce the amount of total solids yogurt (Tamime andRobinson 1999).

No significant differences were observed in the proteinand fat of the different yoghurt samples (Tables 6 and 7).Similarly, Ismail et al. (2006) also reported that there wereno observable differences in TS and F/DM of labnehproduced by addition of six different EOs.

Organoleptic Properties

The mean acceptability scores of the yoghurt samplescontaining different concentrations of EO and probioticbacteria are shown in Table 8. Samples without EO andprobiotic bacteria were used as control groups. The resultsof this study revealed that EO, probiotic culture and theirinteraction have significant (P < 0.05) effect on the orga-noleptic properties of the yoghurt samples. Control sampleand then yoghurt sample contained 40 ppm of EO was themost preferred samples (P < 0.05). In general, the samplescontaining 40, 60 and 80 ppm of the EO with the probioticbacteria were acceptable to panelists.

It can be concluded that 40 ppm of T. polium EO can beused in order to increase the shelf life of yoghurt for up to28 days at 4C with acceptable flavor, good appearance

TABLE 5. CHANGES IN SNF (%) OF YOGHURTSAMPLES DURING THE STORAGEYoghurt

samples


1 7 14 21 28

SNFA 8.20 � 0.09a 8.12 � 0.05a 7.95 � 0.09b 8.00 � 0.03b 7.90 � 0.06b

B 8.09 � 0.13a 7.96 � 0.09b 7.96 � 0.04b 8.00 � 0.06b 8.00 � 0.05b

C 8.12 � 0.05a 8.00 � 0.03b 8.00 � 0.06b 8.02 � 0.09b 8.00 � 0.07b

D 8.21 � 0.03a 8.10 � 0.15a 8.00 � 0.10b 8.01 � 0.05b 7.91 � 0.10b

E 8.10 � 0.08a 8.03 � 0.04a 8.00 � 0.05b 8.00 � 0.05b 7.90 � 0.07b

F 8.13 � 0.10a 8.05 � 0.03a 7.96 � 0.09b 7.96 � 0.07b 7.90 � 0.03b

G 8.21 � 0.03a 8.07 � 0.10a 8.00 � 0.11b 7.98 � 0.10a 7.92 � 0.09b

H 8.14 � 0.06a 8.06 � 0.11a 8.03 � 0.04a 8.00 � 0.03b 7.90 � 0.09b


TABLE 6. CHANGES IN FAT (%) OF YOGHURTSAMPLES DURING STORAGEYoghurt

samples


1 7 14 21 28

FatA 1.78 � 0.08a 1.70 � 0.05a 1.70 � 0.05a 1.70 � 0.03a 1.68 � 0.09a

B 1.89 � 0.03a 1.75 � 0.09a 1.60 � 0.08b 1.60 � 0.12b 1.60 � 0.03b

C 1.90 � 0.05a 1.80 � 0.06a 1.75 � 0.10a 1.75 � 0.05a 1.73 � 0.08a

D 1.89 � 0.09a 1.80 � 0.10a 1.70 � 0.05a 1.70 � 0.03a 1.70 � 0.03a

E 1.81 � 0.10a 1.75 � 0.03a 1.70 � 0.11a 1.67 � 0.08a 1.65 � 0.09a

F 1.82 � 0.11a 1.75 � 0.08a 1.60 � 0.05b 1.60 � 0.10b 1.60 � 0.03b

G 1.89 � 0.05a 1.80 � 0.03a 1.70 � 0.09a 1.67 � 0.05a 1.65 � 0.10a

H 1.81 � 0.09a 1.75 � 0.05a 1.72 � 0.08a 1.70 � 0.03a 1.70 � 0.03a




without any signs of spoilage organisms and also withappropriated viability of L. casei.

Survival of L. casei

The viability of L. casei in bio-yoghurt containing variousconcentration of T. polium EO (40, 60 and 80 ppm) wassignificantly decreased (P < 0.05) after 28 days of storage(Fig. 1). These results are comparable to those obtained forZiziphora clinopodioides on L. acidophilus activity as bio-yoghurt starter culture by Sarabi-Jamab and Niyazmand(2009), mint, thyme and garlic by Sismek et al. (2007) andalso those belong to Ziziphora by Hadad Khodaparast et al.(2007). Different concentrations of herbal EOs can influ-ence the activity of starter bacteria and LAB in fermentativedairy products and this has been investigated by someresearchers (Bayoumi 1992; Hadad Khodaparast et al. 2007;

TABLE 7. CHANGES IN PROTEIN (%) OFYOGHURT SAMPLES DURING STORAGE Yoghurt

samples


1 7 14 21 28

ProteinA 2.95 � 0.09a 2.90 � 0.03a 2.85 � 0.05a 2.81 � 0.07a 2.80 � 0.09a

B 2.93 � 0.05a 2.88 � 0.09a 2.85 � 0.03a 2.82 � 0.03a 2.80 � 0.04a

C 2.86 � 0.08a 2.86 � 0.11a 2.83 � 0.04a 2.83 � 0.09a 2.80 � 0.07a

D 2.90 � 0.03a 2.81 � 0.05a 2.80 � 0.03a 2.78 � 0.05a 2.77 � 0.03a

E 2.92 � 0.10a 2.87 � 0.08a 2.85 � 0.07a 2.82 � 0.08a 2.80 � 0.10a

F 2.96 � 0.07a 2.90 � 0.03a 2.90 � 0.03a 2.86 � 0.03a 2.85 � 0.05a

G 2.90 � 0.05a 2.85 � 0.03a 2.80 � 0.05a 2.77 � 0.09a 2.77 � 0.07a

H 2.93 � 0.04a 2.90 � 0.08a 2.87 � 0.05a 2.85 � 0.04a 2.85 � 0.04a


TABLE 8. ORGANOLEPTIC PROPERTIES OF YOGHURT FORMULATEDWITH T. POLIUM EO AND L. CASEI

Yoghurt samples Mean � SD

A 7.80 � 0.00aB 7.00 � 1.76b

C 5.14 � 1.51c

D 6.02 � 1.46d

E 7.65 � 0.40e

F 4.26 � 1.05f

G 5.06 � 1.63g

H 6.56 � 2.31h

The means value followed by the same letter are not significantly differ-ent (P < 0.05).Sample A, yoghurt with no additive; sample B, probiotic yoghurt;sample C, probiotic yoghurt with 80 ppm EO; sample D, probioticyoghurt with 60 ppm EO; sample E, probiotic yoghurt with 40 ppm EO;sample F, yoghurt with 80 ppm EO; sample G, yoghurt with 60 ppmEO; sample H, yoghurt with 40 ppm EO.

Sample B

9

8

7

6

5

Sample C

Sample D

1 7 14Storage period of yoghurt (day)

Sur

viva

l of L

. cas

ei (

log1

0 cf

u/g)

21 28

Sample E

FIG. 1. SURVIVAL OF L. CASEI (CFU/G)DURING YOGHURT STORAGE PERIOD



Sismek et al. 2007). Among gram-positive bacteria, LAB areoften known as the most resistant species against antimicro-bial agents of herbs (Lemay et al. 2002). In present studywith regarding result of enumeration of the L. casei, sampleB (probiotic yoghurt) and sample D (treatment containing60 ppm of this EO) had the highest (P < 0.05) total viablecount of probiotic bacteria (6.47cfu/g). Yoghurt manufac-tured with 80 ppm of the T. polium EO had the lowest count(Fig. 1).

Based on the results, L. casei populations were not inhib-ited by low concentrations of the T. polium EO. However,increases in the EO concentrations lead to decreases in bac-terial counts (P < 0.05). It has previously been reported thataddition of some EOs to yoghurt and labneh cheese duringits manufacture had a stimulatory effect on LAB by enhanc-ing their growth and acid production (El-Nawawy et al.1998; Khaleel 2000). Notably, El-Nawawy et al. (1998)reported that the presence of some herbs, including thyme,in the manufacture of yoghurt increased the counts ofS. thermophilus and L. bulgaricus compared to untreatedcontrols during storage.

According to the Food and Agriculture Organization(FAO), a standard probiotic product must contain aminimum of 106–107 cfu/g live and active probiotic micro-organisms at the moment of consumption (FAO/WHO2002). Durability of L. casei at the end storage period ofyoghurt in control and yoghurt containing T. polium EO(especially in 60 ppm) was sufficient to exert beneficialhealth effects.

CONCLUSION

Based on our results, yoghurt can be a very suitable foodproduct to carry relevant probiotic bacteria while addingcertain herbal EOs. In this study, T. polium EO had adverseeffect neither on physicochemical properties nor on theviability of L. casei of yoghurt. In addition, T. polium EO isnontoxic and safe for public health, so it can be an accept-able alternative to chemical preservatives.

REFERENCES

ABBAS, F.M. and OSMAN, M.M. 1998. Properties oflabneh like products manufactured using acid andacid-rennet coagulation. Ann. Agric. Sci. Moshtohor. 36,401–411.

ABD-EI FATTAH, S.M., YAHIA HASAN, A.S., BAYOMN, H.M.and EISSA, H.A. 2010. The use of lemongrass extracts asantimicrobial and food additive potential in yoghurt. J. Am.Sci. 6, 582–594.

ABURJAI, T., HUDAIB, M. and CAVRINI, V. 2006.Composition of the essential oil from Jordanian germander(Teucrium polium L.). J. Essent. Oil Res. 18, 97–99.

ADOLFSSON, O., MEYDANI, S.N. and RUSSEL, R.M. 2004.Yoghurt and gut function. Am. J. Clin.Nutr. 80, 245–256.

AL OTAIBI, M. and EL DEMERDASH, H. 2008. Improvementof the quality and shelf life of concentrated yoghurt (labneh)by the addition of some essential oils. Afr. J. Microbiol. Res. 2,156–161.

AL-ADAMANY, E., KHATTAR, M., HADDAD, T. andTOUFEILI, P. 2003. Estimation of shelf-life of concentratedyoghurt by monitoring selected microbiological andphysiochemical changes during storage. LWT 36, 407–414.

AMIRI, H. 2008. Essential oil composition of T. orientale L.subsp. Taylori (Boiss). J. Med. Plants 7, 100–104.

ANDERSSON, H., ASP, N.G., BRUCE, A., ROOS, S.,WADSTROM, T. and WOLD, A. 2001. Health effects ofprobiotics and prebiotics. A literature review on humanstudies. Scand. J. Nutr. 45, 58–75.

ANJUM, R.R. and ZAHOOR, T. 2007. Comparative study ofyoghurt prepared by using local isolated and commercialimported starter culture. J. Res. (Sci.) 18, 35–41.

ASSOCIATION OF OFFICIAL ANALYTICAL CHEMIST(AOAC). 1995. Official Methods of Analysis, 16th Ed., Vol. II,AOAC, Arlington, VA.

ASSOCIATION OF OFFICIAL ANALYTICAL CHEMIST(AOAC) 2000. Official Methods of Analysis, 17th Ed., Methods926.08, 933.05, 935.42 and 991.20, AOAC International,Gaithersburg, MD.

BAHAR, Z.F. and MIRZA, M. 2003. Volatile constituentsof Teucrium flavum L. from Iran. J. Essent. Oil Res. 15,106–107.

BAYOUMI, S. 1992. Bacteriostatic effect of some spices andtheir utilization in the manufacture of yoghurt. Chem.Microbiol. Technol. Lebensm. 14, 21–26.

BURT, S. 2004. Essential oils: Their antibacterial properties andpotential applications in foods rev. Int. J. Food Microbiol. 94,223–253.

BYLUND, G. 1995. Cultures and starter manufacture. Dairyprocessing Handbook, Tetra Pack Processing System AB,S-221, 86, Lund, Sweden.

CAKIR, A.M.E., DURU, M., HARMANDAR, R., CIRIMINNA,S., PASSANNATI, A. and KAZLM, K. 1998. Volatileconstituents of Teucrium polium L. from Turkey. J. Essent. OilRes. 10, 113–115.

CON, A.H., CAKMAKCI, S., CAGLAR, A. and GOKALP, H.Y.1996. Effect of different fruits and storage period onmicrobiological qualities of fruit-flavored yoghurt producedin Turkey. J. Food Prot. 59, 402–406.

DRAUGHON, F.A. 2004. Use of botanicals as biopreservatives infood. Food Technol. 58, 2–20.

EIKANI, M.H., GOODARNIA, L. and MIRZA, M. 1999.Comparison between the essential oil of supercritical carbondioxide extract of Teucrium polium L. J. Essent. Oil Res. 11,470–472.

EL-NAWAWY, M.A., EL-KENANY, Y.M. and EL-GHAFFAR,E.A. 1998. Effect of some herb plants on the use of yoghurtculture. Annals of Agriculture Sci. 7th Conf. Agric. Dev. Res.



Fac. Agric. Ain Shams University of Cairo, Egypt. 15–17December.

EUROPEAN PHARMACOPOEIA. 1983. Part 1, MaisonneuveSA, Sainte Ruffine, p. V.4.5.8.

FAO/WHO. 2002. Guidelines for the Evaluation of Probiotics inFood. A joint publication of FAO and WHO, London,Ontario: Canada.

GUARNER, F. and SHAAFSMA, G.J. 1998. Probiotics. Int. J.Food Microbial. 39, 237–238.

HADAD KHODAPARAST, M.H., MEHRABAN- SANGATASH,M., KARAZHYAN, R., HABIBI- NAJAFI, M.B., BEIRAGHI-TOOSI, S. 2007. Effect of essential oil and extract of Ziziphoraclinopodioides on yoghurt starter culture activity. World Sci. J.2, 194–197.

HADDADIN, J.S.Y. 2005. Kinetic studies and sensory analysis oflactic acid bacteria isolated from white cheese made fromsheep raw milk. Pak. J. Nutr. 4, 78–84.

HASANI, P., YASA, N., VOSOUGH-GANBARI, S.,MOHAMMADIRAD, A., DEHGAN, G. and ABDOLLLAHI,M. 2007. In vivo antioxidant potential of Teucrium polium, ascompared to a-tocopherol. Acta Pharm. 57, 123–129.

IDF. 1991. Yoghurt: Determination of titratable acidity. IDFStandard 150, International Dairy Federation.

ISMAIL, A.M., HARBY, S. and SALEM, A.S. 2006. Production offlavored labneh with extended shelf life. Egypt. J. Dairy Sci.34, 59–68.

IWALKOKUM, B.A. and SHITTU, M.O. 2007. Effect of Hibiscussabdariffa (Calyce) extract on biochemical and organolepticproperties of yoghurt. Pak. J. Nutr. 6, 172–182.

JAIMAND, K., REZA, M.B., SOLTANIPOOR, M.A. andMOZAFFARIAN, V. 2006. Volatile constituents of Teucriumstocksia Boiss. Stocksianum from Iran. J. Essential Oil Res. 18,476–477.

KHALEEL, E.M. 2000. Studies on milk and milk products. MScThesis. Faculty of Agriculture, Zagazig University of Egypt.

KULLISSAR, T., ZILMER, M., MIKELSAAR, M., VIHALEMM,T. and KILK, A. 2002. Two antioxidative lactobacilli strains aspromising probiotics. Int. J. Food Microbiol. 72, 215–224.

LEMAY, M.J., CHOQUETTE, J., DELAQUIS, P.J., GARIEPY, C.,RODRIGUE, N. and SAUCIER, L. 2002. Antimicrobial effectof natural preservatives in a cooked and acidified chickenmeat model. Int. J. Food Microbiol. 78, 217–226.

MC GREGOR, J.U. and WHITE, C.H. 1986. Effect of sweetenerson the quality and acceptability of yoghurt. J. Dairy Sci. 69,698–703.

MCFARLAND, L. 2000. A review of evidences of health claimsfor biotherapeutic agents. Microb. Ecol. Health Dis. 12, 65–76.

MEILGAARD, M., CIVILLE, G.V. and CARR, B.T. 1999. SensoryEvaluation Techniques, CRC Press, Boca Raton, FL.

MUHAMMED, B.F., ABUBAKAR, M.M., ADEGBOLA, T.A. andOYAWOYE, E.O. 2005. Effects of culture concentration and

inoculation temperature on physicochemical, microbialand organoleptic properties of yoghurt. Niger. Food J. 23,156–165.

PHILLIP, S.M., KAILASAPATHY, K. and TRAN, L. 2006.Viability of commercial probiotic cultures (L. acidophillus,Bifidobacterium spp., L. casei, L. paracasei and L. rhamnosus)in cheddar cheese. Int. J. Food Microbiol. 108, 276–280.

SALMINEN, S. 2001. Human studies on probiotics: Aspects ofscientific documentation. Scand. J. Nutr. 45, 8–12.

SALVADOR, A. and FISZMAN, S.M. 2004. Textural and sensorycharacteristics of whole and skimmed flavored set-typeyoghurt during long storage. J. Dairy Sci. 87, 4033–4041.

SARABI-JAMAB, M. and NIYAZMAND, R. 2009. Effect ofessential oil of Mentha piperita and Ziziphora clinopodioideson Lactobacillus acidophilus activity as bioyoghurt starterculture. Am. Eurasian J. Agric. Environ. Sci. 6, 129–131.

SHAH, N.P. 2001. Functional foods, probiotics and prebiotics.Food Technol. 55, 46–53.

SHAHRAKI, M.R., ARAB, M.R., MIRIMOKADDAM, E. andJEY-PALAM, A. 2006. The effect of Teucrium polium on liverfunction, serum lipids and glucose in diabetic male rate. Iran.Biomed J. 11, 65–68.

SHIN, J.G., LEE, J.J., KIM, H.Y. and BAEK, Y.J. 1991. Studies onthe changes in quality and sensory evaluation of stirredyoghurt stored at different temperatures. Korean J. Dairy Sci.13, 148–155.

SINGH, G., KAPOOR, I.P.S. and SINGH, P. 2011. Effect ofvolatile oil and oleoresin of anise on the shelf life of yoghurt.J. Food Process. Preserv. 35, 778–783.

SISMEK, B., SAGDIC, O. and OZCEILIK, S. 2007. Survival ofEscherichia coli O157:H7 during the storage of Ayranproduced with different spices. J. Food Eng. 78,676–680.

SONGISEPP, E., KULLISAAR, T., HUTT, P., ELIAS, P.,BRILENE, T., ZILMER, M. and MIKELSAAR, M. 2004. A newprobiotic cheese with antioxidative and antimicrobial activity.J. Dairy Sci. 87, 2017–2023.

TAMIME, A.Y. and ROBINSON, R.K. 1999. Yoghurt. Science andTechnology, Woodhead Publishing, London,U.K.

WOLFSCHOON, A.F. 1983. The total solid contents of milk, theacidity, pH and viscosity of yoghurt. Reviste Do Institute DeLacticinois Candido Tosez 38, 19–24. (Food Sci. and Technol.Abstracts, 16(10), (1984), 2187).

WOOD, B.J.B. 1992. The Lactic Acid Bacteria in Health andDisease, pp. 151–339, Elsevier Appl. Sci., London, England.

YEGANEHZAD, S., MAZAHERI-TEHRANI, M. and SHAHIDI,F. 2007. Studying microbial, physiochemical and sensoryproperties of directly concentrated probiotic yoghurt. Afr. J.Agric. Res. 2(8), 366–369.

ZARGARI, A. 1994. Medicinal Plants, 6th Ed., Tehran UniversityPublication, Tehran.



Effect of Teucrium polium Essential Oil on the Physicochemical and...

Documents

Transcript of Effect of Teucrium polium Essential Oil on the Physicochemical and...