Palm Sap Research Articles

Screening for gum-producing Lactic acid bacteria in Oil palm (Elaeis guineensis) and raphia palm (Raphia regalis) sap from South-West Nigeria.

Lactic acid bacteria have wide applications in food processing. Lactic acid bacteria produced exopolysaccharides (EPS) which could be used as possible replacer for commercial stabilizer and thickeners produced by nonfood grade bacteria. Seventy‐two samples of Oil and Raphia palm sap were collected in eighteen locations across South‐Western Nigeria and screened for exopolysaccharide production in 6% sucrose agar using streaked plate method. Four hundred EPS‐producing bacteria (EPB) isolated were clustered based on morphological characteristics into two broad groups and preliminary screened for EPS‐producing capacity. Twenty representative of EPB were selected from the broad groups for tentative identification by API 50CHL and 10 high yielding EPB were selected for large‐scale EPS production. Each strain was inoculated into 6% sucrose broth with 3% (v/v) preculture grown overnight in a 1.5 ml flask and incubated at 37°C for 72 hr. The EPSs were purified and freeze‐dried prior to quantification of yields. EPS‐producing bacteria were identified as Leuconostoc lactis, Lactobacillus fermentum, Lactobacillus delbrueckii ssp. lactis, L. delbrueckii ssp. delbrueckii, Lactobacillus acidophilus, Lactobacillus plantarum, Lactobacillus crispatus, and Leuconostoc mesenteroides ssp. mesenteroides/dextranicum. EPS yield ranged from 132–810.75 mg/L and EPS‐producing potential of lactic acid bacteria (LAB) strains ranged; 36% (132–245 mg/L), 36% (250–460 mg/L), and 28% (461–820 mg/L). L. plantarum had the highest EPS yield of 810.75 mg/L whereas L. crispatus had the least yield 242.5 mg/L. These results suggest that majority of LAB in palm wine saps are gum‐producing bacteria. Leuconostoc and Lactobacillus were the most abundant LAB found in this study while L. plantarum could have applications as potential starter cultures for the production of exopolysaccharides (EPS) at industrial level.

Indigenous technology in utilizations and handlings of palmyra palm (Borassus flabellifer L.) sap and its quality from two regions of East Java

Indigenous technology in utilizations and handling of palmyra palm sap were studied in two regions of East Java, i.e at Gunungsari village, Beji, Pasuruan, and at two districts in Sumenep, Madura, which were known as one of palmyra palm plantation centre in East Java. Field observations and interviews were carried out to some farmers that taken by purposive sampling. Result of this research shows that there were differences in utilization and handling of sap between two regions. All of farmers in Gunungsari utilize sap in fresh sap called legen as their produce, and some fermented sap, wine called toak and further fermented as vinegar. On the other side, farmers in two districts in Sumenep, 96.7 % at Pragaan and 100 % at Dungkek process their palmyra palm sap into jaggery or palm sugar, this is due to high production of sap in this region as Sumenep still has high palm population. At Gunungsari, farmers preserve sap by boiling fresh sap after four hours tapping process if they do not need to ferment it to wine or vinegar. In Sumenep, according to sap utilization as palm sugar, they preserve the sap by adding sap a pinch of gum-lack tree (Schlechera oleosa) bark or cashew (Anacardium occidentale) leaves during tapping. Analyses of sap samples from two regions taken in wet season showed that total soluble solid contained were not significantly difference, it ranges from 9.47 until 11.67 brix. Reducing sugar contained in sap from Sumenep was the lowest, i.e 0.67 % compared with those at Gunungsari 2.10% for fresh and 2.04% for boiled sap. Sap preserved by adding gum-lack tree bark had higher total soluble solid and reducing sugar than the boiled one. The tapping, collecting, utilizations and handling of palmyra palm sap in two regions still carried out traditionally, technology innovations are urgently needed in these regions.

ISOLASI DAN AKTIVITAS ANTI MIKROBA BAKTERI ASAM LAKTAT DARI FERMENTASI NIRA KELAPA SAWIT

The aim of the study was to isolate lactic acid bacteria (LAB) from palm oil sap and to investigate the anti microbial activity of LAB against pathogen bacteria, Staphylococcus aureus and Escherichia coli using disc diffusion method. Palm oil sap were enriched using spesific medium MRS (Man Rogosa Sharpe) broth overnight, serial dilution were conducted and the culture were spread on the surface of MRS agar and then incubated for 48 hours. The result showed the total colonies of LAB were 1,4 x 107 CFU/mL. 6 colonies were randomly selected namely EHN1, EHN2, EHN3, EHN4, EHN5 and EHN6. The selected colonies are classified as Gram positive and bacilli in morphology. Antimicrobial activity against pathogenic bacteria was investigated using disc diffusion method. Antimicrobial activity of isolates against E.coli and S.aureus were 12,2 mm; 9mm; 11mm; 11mm; 10,5mm; 12 mm dan 13,5mm; 11mm; 12mm; 11,5mm; 12mm; 12,5mm respectively. from these result it can be concluded that LAB isolated from oil palm sap has potential as probiotic.

Colour changes during the processing of Arenga pinnata (palm) sap into sugar.

This study reports the colour changes using CIELAB measurements (L*a*b* color space) and their correlation with time, temperature, total soluble solids (TSS), and pH during the processing of Arenga pinnata (palm) sap into granulated sugar. As a function of time, the sap became darker, more red, and more yellow indicating a continuous decrease of L* values, and an increase in both a* and b* values. A constant decrease of L* and an increase of a* and b* were particularly observed when TSS of the sap was more than 33.5%. During the process, regression analysis showed that a linear pattern was indicated for total colour difference (ΔE*) with time, whereas a non linear relationship was shown for ΔE* with TSS, temperature and pH. Stirring the syrup changed the L*a*b* values while ΔE* showed a decrease throughout the process. This research will serve as a base for future developments for the manufacture and quality grading of A. pinnata (palm) sugar.

EFFECTS OF NIPAH VIRUS IN TODAY’S WORLD

The effect of Nipah Virus Infection is increasing day by day in today’s scenario and more number of cases are found in various countries. In India it was discovered in Sikkim, Siliguri and West Bengal. It is near borders with China, Bangladesh, Nepal, and Sikkim. The primary pathways of transmission is from bats to people, in Bangladesh its transformed via contamination of raw date palm sap by bats with subsequent consumption by humans and through infection of domestic animals (cattle, pigs, and goats), presumably from consumption of food contaminated with bat saliva or urine with subsequent transmission to people. It is found in both species of humans as well as animals more number of deaths was found in the both spices, hence zoonotics. Laboratory investigations at the time of the outbreak did not show or identify an infectious agent. Approximately half of recognized Nipah cases in Bangladesh developed their disease following person to person transmission of the virus. Efforts to prevent transmission should focus on decreasing bat access to date palm sap and reducing family members' and friends' exposure to infected patients' saliva or body fluids.

Economic Valuation of Nipa Palm (Nypa fruticans Wurmb.) Sap as Bioethanol Material

Nipa palm (Nypa fruticans Wurmb.) widely recognized as one of the biological resources with various aspect of utilization. Nipa palm sap can be processed into bioethanol. Bioethanol also has a quite high selling price, especially for the fulfilment of energy and for fossil fuels substitution. The main purpose of the study was to evaluate the economic value of nipa sap for further processed into bioethanol, in order to improve the income and job opportunity of coastal households. This research was conducted at Sungsang, South Sumatera with focusing on nipa palm sap further processing into bioethanol and its impact on increasing household income. This research was statistically testing the relationship between potential production of bioethanol, average revenue and labour recruitment. Based on the 2013 data, potential production of bioethanol was capable to contribute of nett income about IDR 507,940 to 3,503,280 ha-1 month-1 or IDR 6,095,274 to 42,039,360 ha-1 year-1. Therefore each hectare of nipa palm can generate labour recruitment up to 3.7 person ha-1. This assessment was expected to provide a better description of the importance of local biological resources utilization of the nipa palm, in order to improve community welfare and the efforts of biodiversity conservation on a local scale.

Nipah Virus Infection.

Nipah virus, a paramyxovirus related to Hendra virus, first emerged in Malaysia in 1998. Clinical presentation ranges from asymptomatic infection to fatal encephalitis. Malaysia has had no more cases since 1999, but outbreaks continue to occur in Bangladesh and India. In the Malaysia-Singapore outbreak, transmission occurred primarily through contact with pigs, whereas in Bangladesh and India, it is associated with ingestion of contaminated date palm sap and human-to-human transmission. Bats are the main reservoir for this virus, which can cause disease in humans and animals. There are currently no effective therapeutics, and supportive care and prevention are the mainstays of management.

Livelihood and Rural Tourism Development in Coastal Area North Maluku Province Indonesia

The livelihood of the people who live in the villages in coastal and remote areas can be developed into tourism area based on local wisdom to achieve sustainable tourism. Tunuo village is a village that has a wealth of Natural Resources and the Cultural Resources to support tourism. Nevertheless, Tunuo Village has problems related to Human Resources to support tourism. Meanwhile, the livelihood of Tunuo Village as producers of copra have constraints, namely access to economic capital. The gain from copra sale is erratic and prone to change, it encourages people to seek income from other options to meet their needs. These economic pressures encourage the development of environmentally damaging activities, such as fishing using homemade bombs so that reefs may be damaged. There are also other activities, such as the production, distribution, and consumption of traditional liquor made from palm tree sap, locally known as Saguer. Therefore, the present tourism as a sector that could provide opportunities for the villagers of Tunuo to earn extra income as well as efforts to preserve the environment, and culture. This research was conducted in the village of Tunuo, North Kao District, North Halmahera Regency, North Maluku province, Indonesia. The method is qualitative Participatory Action Research (PAR). The results indicate that people have a desire and a strong motivation to develop, so that the process of forming the Tourism Awareness Group (Pokdarwis) and its seven charms (saptapesona) can take place properly. As of the socialization of tourism awareness, as well as capacity building seminar for Pokdarwiswould be able to mobilize the community to clean up trash on the beach and to encourage communities to develop local knowledge into a leading tourism product.

Palm Sap—Quality Profiles, Fermentation Chemistry, and Preservation Methods

Palm sap is quite nutritious and highly prone to fermentation. The unfermented juice could be an ideal health drink. Palm sap’s quality profile and fermentation chemistry help to predict its shelf life and potential safety. There is demand from farmer–producer companies and food processing industries to develop bottling technology and a transportation/distribution protocol for palm sap similar to common soft drinks. Different techniques were followed for bottling palm sap, but none proved successful at the pilot level or commercial scale. To develop a systemic preservation technique, it is crucial to understand the biochemical composition, fermentation chemistry, and existing preservation methods and their disadvantages. This review mainly focuses on the chemical, microbial, volatiles, and flavor changes of palm sap. In addition to a detailed discussion on contemporary sap preservation techniques, this paper also addresses the effect of pasteurization in combination with preservatives, such as nisin, sodium benzoate, chitosan, potassium sorbate, sorbic acid, citric acid, and sodium metabisulfite, on the shelf life of sap, challenges to preserving palm sap, and future directions for preservation methods.

Ultrafiltration and thermal processing effects on Maillard reaction products and biological properties of date palm sap syrups (Phoenix dactylifera L.)

The effect of ultrafiltration process and temperature concentration on MRPs content and antioxidant, antimicrobial and cytotoxic properties of date palm sap syrups were investigated. MRPs were analyzed by HPLC. Antioxidant activity was evaluated by reducing power and DPPH free radical and H2O2 scavenging activities. Antimicrobial activity was evaluated by the agar disk diffusion method. In vitro cytotoxic activity was examined by cell proliferation assay. Date sap syrups displayed strong antioxidant activities which are correlated 5HMF and 2F contents. In addition, concentration at 100 °C, unlike ultrafiltration process, enhanced significantly the antioxidant activities sap syrups and total phenolic contents. The antimicrobial activities showed marked activity against S. enterica, P. aeruginosa, S. aureus, L. monocytogenes with an inhibition zone of 21, 34, 27 and 34 mm respectively. Cytotoxicity assays showed that sap syrups can inhibit the proliferation of HeLa cell lines at high concentration.

ANALISIS KELAYAKAN FINANSIAL DAN STRATEGI PENGEMBANGAN USAHA INDUSTRI RUMAHAN GULA SEMUT (PALM SUGAR) DARI NIRA NIPAH DI KELURAHAN PALLANTIKANG

This research is aim to know a work-development based on the supplying of nira nipah (sap of nipa palm) as a basic ingredient, production, labor, marketing, financial-feasibility, and work-development strategy. The subject of the study were 30 sap-incisor farmers and a palm sugar home industry in Pallantikang village. The feasibility analysis used were Benefit Cost Ration (BCR), Net Present Value (NPV), Internal Rate of Return (IRR), Break Event Point (BEP), and PBP (Pay Back Period) to know the feasibility of the work. SWOT matrices was used to build work-development strategy of the palm sugar home industry. The result of the study showed that the work-development strategy of a basic ingredient, production, labor, and marketing are suitably done in the research location. The financial-feasibility toward palm sugar home industry in Pallantikang village for the BCR, IRR, BEP (Unit), BEP (Rupiah), and PBP criterion is suitably done. Moreover, based on SWOT analysis, it is found a recommendation for this research that are increasing the operating income; work in cooperation with government to get a capital aid, tools, even training for the owner and the labors of palm sugar, and give a development and training for sap-incisors in order to not making ballo or fermented palm wine

Aroma profile and appearance of dark chocolate formulated with palm sugar–sucrose blends

Palm sap sugar is a natural alternative sweetener that can be made from the sap/nectar of several species of palm tree flowers. In this work, a thorough investigation about the impact of chemical composition and physical properties of palm sugar on the formation of aroma and appearance of dark chocolate was carried out. Five sucrose–palm sugar blends with different palm sugar (PS) proportion, namely PS0, PS25, PS50, PS75, and PS100, were used as sweetener. The results showed that a higher concentration of some alkyl pyrazines, alkyl furans, alkyl alcohols, 2-acetylpyrrole, and acetic acid observed in the palm sugar-sweetened dark chocolate could be attributed to the aroma profile of palm sugar. In addition, there were some aroma volatiles which were only observed in palm sugar-sweetened dark chocolate, such as 2-furoic acid, 3-hydroxy-2-butanone, 1-hydroxy-2-propanone, 2,3-dihydro-5-dihydroxy-6-methyl-4H-pyran-4-one, and 2,3-dihydro-3,5-dihydroxy-6-methyl-4H-pyran-4-one (DDMP). The higher the proportion of palm sugar, the higher the concentration of these volatiles. Aside from this, the presence of more amorphous sugar was probably responsible for a higher concentration of aroma volatiles in chocolates containing palm sugar. Regarding the chocolate appearance, the effect of particle size in determining the colour was less pronounced than the effect of particle density and possible sugar network formation in the chocolate. The latter factor, might affect the surface roughness, resulting in different colour parameters. It seemed that the Maillard reaction during chocolate processing, which may influence the aroma profile and darkness of chocolate, occurred in much lesser extent that in the palm sugar production.

Epidemiology and factors associated with amoebic liver abscess in northern Sri Lanka

BackgroundClinically diagnosed amoebic liver abscess (ALA) caused by Entamoeba histolytica has been an important public health problem in Jaffna district, northern Sri Lanka for last three decades. In order to draw up a control strategy for elimination of this condition, knowledge of its epidemiology and factors associated with this condition in the local context is vital.MethodsAll clinically diagnosed ALA patients admitted to the Teaching Hospital, Jaffna during the study period were included in the study and the data were collected using an interviewer administered questionnaire. One hundred blood samples from randomly selected toddy (a local alcoholic drink consisting of the fermented sap of the Palmyrah palm) consumers and 200 toddy samples were collected. Toddy samples were cultured in Robinson’s medium to establish the presence of Entamoeba histolytica in the sample. Climatic data and the total toddy sales in the district were obtained from the Meteorological and Excise Departments respectively. A sub group of randomly selected 100 patients were compared with 100 toddy consumers who were negative for E. histolytica antibody to explore the potential risk factors.ResultsBetween July 2012 and July 2015, 346 of 367 ALA patients were enrolled in this study. Almost all patients (98.6%) were males with a history of heavy consumption of alcohol (100%). Almost all (94.2%) were within the age group 31–50 years. None of the cultured toddy samples grew E. histolytica. The monthly incidence of disease peaked in the dry season, matching the total toddy sales in the district. Age, type of alcohol and frequency of drinking were identified as potential risk factors whereas frequency of alcohol consumption and type of alcohol (consuming toddy and arrack) were identified as the independent risk factors. Moreover, the knowledge, attitude and practices towards ALA were poor among participants and the control group.ConclusionsThough the number of cases has declined in recent years, ALA still remains as an important public health problem in Jaffna district. The transmission route of E. histolytica leading to ALA has to be further explored. Moreover, greater awareness among the public who are at risk would be beneficial in order to eliminate the disease.

Review on the study of health effects of some fermented foods consumed on Côte d'Ivoire

Traditionally produced starchy foods and fermented beverages contribute to the diet of people. The benefits of these fermented traditional foods for and their nutritional quality are poorly known. This review gives a synoptic view of some fermented foods consumed in Cote d'Ivoire. Several foods are fermented traditionally, including cereals (maize, millet and sorghum), tubers (cassava), fruits (cocoa beans), palm sap (bandji) and fish (Galeoides decadactylus, Oreochromis niloticus). These fermented foods have varying qualities. Lactic acid bacteria are the main microorganisms of their fermentations. Beyond their nutritional quality, fermented foods and fermentation actors may have other health effects. The survival capacity of lactic acid bacteria at the conditions encountered in the digestive tract and their effect on the maintenance of the integrity of the digestive tract are particularly interesting. This review focuses on the benefits of fermented foods including nutritional aspects, benefits and lactic microflora that occurs during spontaneous fermentations as well as their probiotic effects will also be examined.

Production and structural characterization of exopolysaccharides from newly isolated probiotic lactic acid bacteria

In this work, four exopolysaccharide-producing lactic acid bacteria (LAB) strains, newly isolated from Tunisian spontaneously fermented foods and beverages, namely bovine and turkey meat sausages (BMS and TMS), date palm sap (DPS) and cow milk (CM), were identified as Leuconostoc citreum-BMS, Leuconostoc mesenteroides-TMS, Pediococcus pentosaceus-DPS and Leuconostoc pseudomesenteroides-CM, respectively. The isolated strains showed the ability to withstand simulated human gastrointestinal (GI) tract conditions (low pH, lysozyme, bile salts, pepsin and pancreatin) and showed high surface hydrophobicity (79–90%), besides their ability to act against Escherichia coli and Listeria monocytogenes and to produce exopolysaccharides (EPS). Therefore, these isolates can be served as potential probiotics. The produced EPS were growth-associated suggesting that they are primary metabolites. The molecular weights were higher than 106Da using HPLC-SEC. 2D-NMR results indicated that all the samples were mixtures of dextran and levan, except for EPS-CM which was a levan-type EPS. Furthermore, the EPS samples showed an abitlity to inhibit and to disrupt pathogenic biofilms and showed high thermostability studied via differential scanning calorimetry (DSC) with melting points higher than 224°C making them promising to be used in thermal processed foods.

Chemical Composition of Some Natural Palm Wine Preservatives

Palm wine is the commonest name of the beverage obtained from fermented palm sap, the exudate from tapped unopened spathe of oil palm tree (Elaeis guineensis). This refreshing wine of West and Central Africa is very sweet but within 24 hours the concentration of sucrose falls to less than 50% the initial amounts due to a rapid sugar fermentation by microorganisms. In Cameroon, Central Africa, traditional attempts to preserve palm wine imply the introduction of natural preservatives mostly barks and leaves from edible or medicinal plants. The leaves of Cymbopogon citratus, Pimenta racemosa, Vernonia amygdalina, Ocimum basilicum and the barks and the leaves of Garcinia lucida and of Adansonia digitata commonly used in palm wine preservation were analyzed for their chemical composition. The results showed that lipid, protein, ash, crude fiber, total sugars and vitamin C contents were respectively from 15.70 ± 1.13 to 23.57 ± 1.80%, from 8.38 ± 0.38 to 43.81 ± 3.69%, from 1.14 ± 0.17 to 10.06 ± 0.39%, from 11.74 ± 1.11 to 48.42 ± 0.55%, and from 1.49 ± 0.02 to 22.56 ± 0.59% and from 55.76 ± 3.15 to 175.71 ± 2.09 μg /100 g dw. The phyto-chemicals levels were respectively from 1.37 ± 0.22 to 3.61 ± 0.6 % for total alkaloids, 2.66 ± 0.12 to 4.80 ± 0.24 % for total phenolics and 0.57 ± 0.09 to 1.88 ± 0.09 mg /100 g dw for total saponins. Concerning anti-nutrients, the oxalates levels were higher than the threshold value (250 mg/100 g) reported as safety limit. Despise the proven antimicrobial benefits of alkaloids, saponins and phenolics found in those leaves and barks, caution shall be paid during their use due to their high oxalates levels.

Study of Fermentation Kinetics of Palm Sap from Cocos nucifera

Palm wine plays an important role as an alcoholic beverage in traditional practices. It is important to study the biochemical characteristics and microbiological aspects to understand the fermentation kinetics of palm saps. In the present investigation an elaborate study was carried out to study the fermentation kinetics of coconut palm sap. Total sugar, reducing sugar content and glucose concentration was estimated periodically during fermentation for 16h. Microbial load and invertase assay results were related to the changes in sugar concentration. Initial predominance of lactic acid bacteria was followed by dominance of yeasts. Hydrolysis of non reducing sugar occured at a faster rate between 3-9h of fermentation. During this period, the multiplication of yeasts began and reached its peak at 11h fermentation. Ethanol concentration was around 4.0 and 4.1% at 11h and 13h of fermentation respectively.Int. J. Appl. Sci. Biotechnol. Vol 5(3): 375-381

Convergence of Humans, Bats, Trees, and Culture in Nipah Virus Transmission, Bangladesh.

Preventing emergence of new zoonotic viruses depends on understanding determinants for human risk. Nipah virus (NiV) is a lethal zoonotic pathogen that has spilled over from bats into human populations, with limited person-to-person transmission. We examined ecologic and human behavioral drivers of geographic variation for risk of NiV infection in Bangladesh. We visited 60 villages during 2011–2013 where cases of infection with NiV were identified and 147 control villages. We compared case villages with control villages for most likely drivers for risk of infection, including number of bats, persons, and date palm sap trees, and human date palm sap consumption behavior. Case villages were similar to control villages in many ways, including number of bats, persons, and date palm sap trees, but had a higher proportion of households in which someone drank sap. Reducing human consumption of sap could reduce virus transmission and risk for emergence of a more highly transmissible NiV strain.

A VLP-based vaccine provides complete protection against Nipah virus challenge following multiple-dose or single-dose vaccination schedules in a hamster model

Nipah virus is a highly lethal zoonotic paramyxovirus that was first recognized in Malaysia during an outbreak in 1998. During this outbreak, Nipah virus infection caused a severe febrile neurological disease in humans who worked in close contact with infected pigs. The case fatality rate in humans was approximately 40%. Since 2001, NiV has re-emerged in Bangladesh and India where fruit bats (Pteropus spp.) have been identified as the principal reservoir of the virus. Transmission to humans is considered to be bat-to-human via food contaminated with bat saliva, or consumption of contaminated raw date palm sap, although human-to-human transmission of Nipah virus has also been documented. To date, there are no approved prophylactic options or treatment for NiV infection. In this study, we produced mammalian cell-derived native Nipah virus-like particles composed of Nipah virus G, F and M proteins for use as a novel Nipah virus vaccine. Previous studies demonstrated that the virus-like particles were structurally similar to authentic virus, functionally assembled and immunoreactive. In the studies reported here, purified Nipah virus-like particles were utilized either alone or with adjuvant to vaccinate golden Syrian hamsters with either three-dose or one-dose vaccination regimens followed by virus challenge. These studies found that Nipah virus-like particle immunization of hamsters induced significant neutralizing antibody titers and provided complete protection to all vaccinated animals following either single or three-dose vaccine schedules. These studies prove the feasibility of a virus-like particle-based vaccine for protection against Nipah virus infection.

Comparative Evaluation of the Methods of Palm Sap Fermentation for Bio-ethanol Production

This paper evaluates the methods of fermenting palm sap for bio ethanol production and compares the yield. A survey was carried out in three local government in Akwa Ibom State, Nigeria; Essien Udim, Ikot Abasi and Uruan local government area. Personal interviews were conducted with local palm wine dealers and questionnaires distributed. Three types of palm trees were found dominant in these areas; oil, raphia and nypa palm tree. The sap from these three palm trees were used in investigation. Two methods were investigated; natural and yeasted fermentation using saccharomyces cerevisae. 60% respondents of the questionnaires agreed on using additives to speed up fermentation. Experiments were carried out in which the result shows that yeasted fermentation yields more alcohol than natural fermentation and also reduce the period of fermentation.