Thesis (PhD Food Sc)--Stellenbosch University, 2012. / ENGLISH ABSTRACT: The fermentation of milk has been known for millennia and leads to nutritious and prolonged shelf-life dairy products. In Southern Africa, traditional fermented dairy products have the same value as local staple foods and are consumed as a part of or as a whole meal. However, the retail price and the technology make many commercialised fermented dairy products unaffordable to the majority of the population. There is thus a need for a healthy nutritious low-cost easily prepared fermented dairy product. A product that could be the answer to the above need, is Kefir. The principle advantage is that the Kefir beverage is made from reusable Kefir grains, which unfortunately are not easily available and grow slowly. Kefir grains can only be obtained from pre-existing grains, which presents a problem in the marketing of the grains. A mass culturing technique was developed to produce large masses of grains but preparation of Kefir using these grains results in a product (MG Kefir) lacking in the sensory attributes of Traditional Kefir. Thus, the overall objective of this research was to determine the impact of environmental factors on the metabolic profiles of Kefir produced using different Kefir grains and this was then followed by the subsequent enrichment of Kefir prepared with mass cultured grains so as to obtain a Kefir beverage that has improved organoleptic qualities. To determine the impact of environmental factors Traditional and MG Kefir were prepared under controlled and uncontrolled conditions. Traditional Kefir was found to give the best beverage and was thus considered as the control. Under controlled conditions the optimum incubation temperature for the production of Kefir was 22ºC as over-acidification was observed at 25ºC. The metabolic profiles of both Traditional and MG Kefir showed that both contained acetaldehyde, ethanol, acetone, diacetyl and acetic acid. In addition, the metabolic profiles revealed that an inadequate ratio of diacetyl to acetaldehyde as well as the lack of ethyl acetate was responsible for the flavour defect in MG Kefir. In order to overcome this defect, citrate and ascorbate (0.015 % w.v-1) were added during Kefir fermentation to enhance the diacetyl and ethyl acetate production. This addition showed a positive impact on diacetyl but not on ethyl acetate production. Improvement of the overall flavour of Kefir was observed as the ratios of diacetyl to acetaldehyde were higher (0.21 – 0.5) in the samples with added citrate and ascorbate than in the samples without (0.12 – 0.17). The production of ethyl acetate in MG Kefir was enhanced by combining the effects of longer incubation (24 h + 18 h at 22ºC), addition of ethanol and acetic acid at 0.79% (m.v-1) and the addition of either Lactococcus lactis ssp. diacetylactis biovar diacetylactis 318 or Candida kefyr 1283. The best yields were obtained in samples containing C. kefyr 1283 and only added ethanol (9.22 mg.L-1), indicating that ethanol is an important factor in ethyl acetate production by Kefir starter strains and suggesting that the absence of ethyl acetate is an indication that the grains do not contain a yeast capable of producing sufficient ethyl acetate. During this investigation, the impact of ethyl acetate on the organoleptic quality of Kefir during storage at refrigerated and room temperatures were also studied. The results indicated that refrigerated Kefir contained up to 40 mg.L-1 of ethyl acetate and was not found defective and thus ethyl acetate was considered a positive contributor to Kefir flavour. This is of particular interest as ethyl acetate is a potent flavour compound at concentrations below 5 mg.L-1. Improvements of MG Kefir’s flavour were successful and will be of value for commercial Kefir production where the main aim is to optimise the flavour of Kefir. However, stabilising the grain microbial consortium was found to be important as it is responsible, over time, for both stable and acceptable Kefir. Acceptability of Traditional, MG and other Kefirs (Candi-Kefir and Lacto-Kefir) prepared with microbially stabilised MG was evaluated by 85 consumers. Results indicated that pH (r = 0.978; p < 0.05) was a significant driver of liking for flavour, especially for female consumers (r = 0.982; p < 0.05). In addition, three clusters, each characterised by different liking attributes were identified. Cluster I generally disliked all the products whereas slight acidic Kefir such as Candi-Kefir (7.63) and Lacto-Kefir (7.09) were preferred by Cluster III. Cluster II showed preference to Kefir with moderate acidity and high ethanol content. In that regard, Traditional Kefir obtained the best score (7.50) and MG Kefir the lowest score (4.87). The sensory study is of value as it led to the identification of the drivers of consumers liking by the different types of consumers. In the course of this project, near infrared reflectance spectroscopy was developed as a rapid method to estimate lactic and acetic acids, which are the organic acids responsible for acidity in Kefir, as well as pH and titratable acidity (TA). The results showed that the calibration models for lactic acid (RPD = 2.57 – 3.16), pH (RPD = 2.90) and TA (RPD = 2.60) were good for screening purposes (2 < RPD < 3); indicating that these models would show if the concentrations of lactic acid, the pH or the TA varied from the normal range. This study has demonstrated that the flavour of MG Kefir, prepared with enriched grains, was successfully improved and has provided some understanding on the preference liking of Kefir, an unknown fermented dairy product to South African consumers. / AFRIKAANSE OPSOMMING: Die fermentering van melk is al vir millennia bekend en lei tot voedsame suiwelprodukte met 'n verlengde raklewe. In Suidelike Afrika het tradisioneel gefermenteerde suiwelprodukte dieselfde waarde as plaaslike stapelvoedsels en word dit as 'n maaltyd of as deel van 'n maaltyd geëet. Die kleinhandelsprys en tegnologie van kommersieel gefermenteerde suiwelprodukte maak hierdie produkte egter onbekostigbaar vir die grootste deel van die populasie. Daar is dus 'n behoefte aan 'n gesonde, voedsame, goedkoop, maklik-om-te-berei gefermenteerde suiwelproduk. 'n Moontlike produk om aan die bogenoemde te voldoen is Kefir. Die hoof voordeel is dat die Kefir drankie van herbruikbare Kefirkorrels gemaak word, maar ongelukkig is hierdie korrels nie vrylik beskikbaar nie, en vermeerder dit stadig. Kefirkorrels kan net van reeds bestaande korrels verkry word wat problematies is vir die bemarking van hierdie korrels. 'n Massakwekingstegniek is ontwikkel vir die produksie van groot hoeveelhede korrels maar die voorbereiding van Kefir met hierdie korrels lei tot 'n produk (MG Kefir) wat sensories minder aanvaarbaar is as tradisionele Kefir. Die hoofdoel van hierdie navorsing was dus om die invloed van omgewingsfaktore op die metaboliese profiele van Kefir, berei deur gebruik te maak van verskillende Kefirkorrels, te bepaal. Dit is gevolg deur die verryking van Kefir berei van massagekweekte korrels om 'n Kefir drankie met verbeterde organoleptiese kwaliteite te verkry. Tradisionele en MG Kefir is voorberei onder gekontroleerde en ongekontroleerde toestande om die impak van omgewingsfaktore te bepaal. Die beste drankie is van tradisionele Kefir verkry en is dus beskou as die kontrole. Die optimum temperatuur vir die produksie van Kefir onder gekonroleerde toestande is 22ºC aangesien oor-versuring by 25ºC waargeneem is. Die metaboliese profiele van beide tradisionele en MG Kefir het gewys dat beide produkte asetaldehied, etanol, asetoon, diasetiel en asynsuur bevat. Die metaboliese profiele het verder gewys dat 'n onvoldoende diasetiel tot asetaldehied verhouding sowel as 'n tekort aan etielasetaat verantwoordelik was vir 'n geur defek in MG Kefir. Om hierdie defek te oorkom is sitraat en askorbaat (0.015% m.v-1) tydens Kefir fermentasie bygevoeg om diasetiel en etielasetaat produksie te verhoog. Hierdie byvoeging het 'n positiewe effek gehad op diasetiel produksie, maar nie op die produksie etielasetaat nie. 'n Verbetering in die algehele geur van Kefir is waargeneem aangesien die diasetiel tot asetaldehied verhoudings hoër (0.21 – 0.5) was in die monsters met bygevoegde sitraat en askorbaat as in die monsters daarsonder (0.12 – 0.17). Die produksie van etielasetaat in MG Kefir is verhoog deur die effekte van 'n verlengde inkubasie tydperk (24 h + 18 h by 22ºC), die byvoeging van etanol en asynsuur teen 0.79% (m.v-1) en die byvoeging van óf Lactococcus lactis ssp. diacetylactis biovar diacetylactis 318 óf Candida kefyr 1283 te kombineer. Die beste opbrengs is verkry in monsters wat C. kefyr 1283 en slegs etanol (9.22 mg.L-1) bevat het. Dit dui daarop dat etanol 'n belangrike faktor is vir etielasetaat produksie in Kefir beginstamme en wys moontlik op die afwesigheid van etielasetaat wat daarop dui dat die korrels nie 'n gis bevat wat bevoeg is om genoegsame hoeveelhede etielasetaat te produseer nie. Tydens hierdie ondersoek is die impak van etielasetaat op die organoleptiese kwaliteit van Kefir gedurende opberging by verkoelde- en kamertemperatuur ook bestudeer. Die resultate het gewys dat verkoelde Kefir tot 40 mg.L-1 etielasetaat bevat het sonder dat dit defektief was. Etielasetaat is dus beskou as 'n positiewe bydraer in terme van Kefir geur. Dit is van besondere belang aangesien etielasetaat 'n sterk geurkomponent teen konsentrasies laer as 5 mg.L-1 is. Verbeteringe in MG Kefir se geur was suksesvol en sal van waarde wees vir kommersiële Kefir produksie waar die hoofdoel die optimisering van Kefir geur is. Stabilisering van die korrels se mikrobiologiese konsortium is ook belangrik aangesien daar gevind is dat dit oor tyd verantwoordelik is vir stabiele en aanvaarbare Kefir. Die aanvaarbaarheid van tradisioneel, MG en ander Kefirs (Candi-Kefir en Lacto-Kefir), voorberei van mikrobiologies gestabiliseerde MG, is deur 85 verbruikers geëvalueer. Die resultate het aangedui dat pH (r = 0.978; p < 0.05) 'n belangrike faktor is in die bepaling van verbruikers se voorkeur van geur is, veral by vroulike verbruikers (r = 0.978; p < 0.05). Drie groepe, elk gekenmerk deur verskillende voorkeur en aanvaarbaarheid eienskappe, is verder geïdentifiseer. Groep I het oor die algemeen van geen van die produkte gehou nie en Groep III het die effense suur Kefirs soos Candi-Kefir (7.63) en Lacto-Kefir (7.09) verkies. Groep II het die Kefir met 'n matige suurheid en hoë etanolinhoud verkies. Tradisionele Kefir het die hoogste telling (7.50) en MG Kefir die laagste telling (4.78) behaal. Die sensoriese studie is van waarde aangesien dit gelei het tot die identifisering van die drywers van verbruikersvoorkeure deur die verskillende tipes verbruikers. Tydens hierdie projek is 'n naby-infrarooi reflektansie spektroskopiese metode ontwikkel vir die vinnige skatting van melk- en asynsuur, die organise sure wat verantwoordelik is vir die suurheid van Kefir, asook die pH en titreerbare suurheid (TS). Die resultate het getoon dat die kalibrasiemodelle vir melksuur (RPD = 2.57 – 3.16), pH (RPD = 2.90) en TS (RPD = 2.60) voldoende was vir siftingsdoeleindes (2 < RPD < 3). Dit dui daarop dat hierdie modelle sal aandui wanneer die konsentrasie van melksuur, pH of TS afwissel van die normale reeks. Hierdie studie het gewys dat die geur van MG Kefir, berei van verrykte korrels, suksesvol verbeter is en het ook gelei tot insigte in die voorkeur van aanvaarbaarheid van Kefir, 'n onbekende gefermenteerde suiwelproduk vir Suid-Afrikaanse verbruikers.