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Polyphasic approach to the identification and characterization of aflatoxigenic strains of Aspergillus section Flavi isolated from peanuts and peanut-based products marketed in Malaysia

Norlia M, Jinap S, Nor-Khaizura MAR, Son R, Chin CK, Sardjono

Int J Food Microbiol, 2018 Oct 03;282:9-15.
PMID: 29885975 DOI: 10.1016/j.ijfoodmicro.2018.05.030

Peanuts are widely consumed as the main ingredient in many local dishes in Malaysia. However, the tropical climate in Malaysia (high temperature and humidity) favours the growth of fungi from Aspergillus section Flavi, especially during storage. Most of the species from this section, such as A. flavus, A. parasiticus and A. nomius, are natural producers of aflatoxins. Precise identification of local isolates and information regarding their ability to produce aflatoxins are very important to evaluate the safety of food marketed in Malaysia. Therefore, this study aimed to identify and characterize the aflatoxigenic and non-aflatoxigenic strains of Aspergillus section Flavi in peanuts and peanut-based products. A polyphasic approach, consisting of morphological and chemical characterizations was applied to 128 isolates originating from raw peanuts and peanut-based products. On the basis of morphological characters, 127 positively identified as Aspergillus flavus, and the other as A. nomius. Chemical characterization revealed six chemotype profiles which indicates diversity of toxigenic potential. About 58.6%, 68.5%, and 100% of the isolates are positive for aflatoxins, cyclopiazonic acid and aspergillic acid productions respectively. The majority of the isolates originating from raw peanut samples (64.8%) were aflatoxigenic, while those from peanut-based products were less toxigenic (39.1%). The precise identification of these species may help in developing control strategies for aflatoxigenic fungi and aflatoxin contamination in peanuts, especially during storage. These findings also highlight the possibility of the co-occurrence of other toxins, which could increase the potential toxic effects of peanuts.
Reduction of Aspergillus spp. and aflatoxins in peanut sauce processing by oil-less frying of chilli powder and retort processing

Farawahida AH, Jinap S, Nor-Khaizura MAR, Samsudin NIP

Food Addit Contam Part A Chem Anal Control Expo Risk Assess, 2017 Dec;34(12):2242-2250.
PMID: 28871861 DOI: 10.1080/19440049.2017.1375605

Among the many roles played by small and medium enterprises (SMEs) in the food industry is the production of heritage foods such as peanut sauce. Unfortunately, the safety of peanut sauce is not always assured as the processing line is not controlled. Peanut sauce is usually made of peanuts and chilli, and these commodities are normally contaminated with Aspergillus spp. and aflatoxins (AFs). Hence, the objective of this study was to evaluate the practices related to reduction of AF hazard and the effect of interventions in peanut sauce processing. Peanut samples were collected from each step of peanut sauce processing from a small peanut sauce company according to four designs: (1) control; (2) oil-less frying of chilli powder; (3) addition of retort processing; and (4) combination of oil-less frying of chilli powder and retort processing. Oil-less frying of chilli powder (Design 2) reduced total AFs by 33-41%, retort processing (Design 3) reduced total AFs by 49%, while combination of these two thermal processes (Design 4) significantly reduced total AFs, by 57%. The present work demonstrated that Design 4 yielded the highest reduction of total AFs and is therefore recommended to be employed by SME companies.
Effects of plasticizers on the physicochemical properties of kappa-carrageenan films extracted from Eucheuma cottonii

Ili Balqis AM, Nor Khaizura MAR, Russly AR, Nur Hanani ZA

Int J Biol Macromol, 2017 Oct;103:721-732.
PMID: 28528954 DOI: 10.1016/j.ijbiomac.2017.05.105

The physicochemical properties of κ-carrageenan films extracted from Eucheuma cottonii (E. cottonii) incorporated with different concentrations and types of plasticizers were studied. Glycerol, sorbitol, and polyethylene glycol-300 (PEG-300) in the range of 10-60% were used as plasticizers. The results showed that the thickness and moisture content (MC) of films increased significantly (p≤0.05) with the increase in plasticizer concentration. Sorbitol-plasticized films had the lowest values. Sorbitol-plasticized films have better mechanical properties and the lowest water vapor permeability (WVP), solubility and water uptake ratio (WUR) compared with glycerol and PEG-plasticized films (p≤0.05). Fourier transform infrared (FTIR) spectra showed the intermolecular reactions between κ-carrageenan and the plasticizers in the films. Scanning electron microscopy (SEM) observations indicated that sorbitol-plasticized films have a compact structure, even at the highest concentration. The melting temperature (Tm) of films decreased (p≤0.05) with an increase in the plasticizer concentration. Here, the glycerol-plasticized films had the lowest values. X-ray diffraction (XRD) showed broad and narrow peaks of the un-plasticized κ-carrageenan film at 2θ=20.0° and 2θ=8.4°, respectively. The intensity of the broad peak increased and the narrow peak disappeared as the concentration of plasticizers increased. In conclusion, films from E. cottonii successfully produced with sorbitol as the plasticizer exhibited good physical properties as packaging films.
Evaluation of aflatoxin and Aspergillus sp. contamination in raw peanuts and peanut-based products along this supply chain in Malaysia

Norlia M, Nor-Khaizura MAR, Selamat J, Abu Bakar F, Radu S, Chin CK

Food Addit Contam Part A Chem Anal Control Expo Risk Assess, 2018 Sep;35(9):1787-1802.
PMID: 29912639 DOI: 10.1080/19440049.2018.1488276

The peanut supply chain in Malaysia is dominated by three main stakeholders (importers, manufacturers, retailers). The present study aimed to determine the levels and critical points of aflatoxin and fungal contamination in peanuts along the supply chain. Specifically, two types of raw peanuts and six types of peanut-based products were collected (N = 178). Samples were analysed for aflatoxins by using high-performance liquid chromatography. Results revealed that the aflatoxin contamination was significantly higher (P ≤ 0.05) in raw peanuts and peanut-based products from the retailers. However, there was no significant difference (P ≥ 0.05) in fungal contamination for both types of peanuts except for the total fungal count in raw peanuts from the retailers. Furthermore, raw peanut kernels from the retailers were the most contaminated ones ranged from
Fulltext Microbiological and Sensorial Quality of Beef Meat (Longissimus dorsi) Marinated with Cinnamon Extract and Stored at Various Temperatures

Zawani CJ, Nor-Khaizura MAR, Mahyudin NA, Ismail-Fitry MR, Nirmal NP

Foods, 2022 Dec 08;11(24).
PMID: 36553712 DOI: 10.3390/foods11243971

Meat spoilage caused by temperature abuse is a major problem for producers, retailers, and consumers that can generate large economic losses to industries. Microbial growth of Pseudomonas spp. is the main source of spoilage during storage. Cinnamon has antimicrobial properties that may potentially be used to reduce the spoilage caused by Pseudomonas. The objectives of this study were to determine the inhibitory effect of cinnamon extract (CE) against Pseudomonas aeruginosa (ATCC 27853) and evaluate the treatment of CE on meat quality during different storage temperatures (5 °C, 10 °C, 15 °C, and 25 °C). The anti-Pseudomonas result showed that 100% (w/v) CE concentration produced a 13.50 mm zone of inhibition in a disc diffusion assay. The minimum inhibitor concentration (MIC) of CE was noted at 25% (v/v), whereas the minimum bactericidal concentration (MBC) value was observed at 50% (v/v) concentration of CE. The time-kill showed the growth of P. aeruginosa decreased from 7.64 to 5.39 log CFU/mL at MIC concentration. Total phenolic content and IC50 value of the cinnamon extract was expressed as 6.72 ± 0.87 mg GAE/g extract and 0.15 mg/mL, respectively. When the meat was marinated with 50% (v/v) CE and stored at various temperatures, the total viable count (TVC) and growth of Pseudomonas spp. were lowered as compared to the control sample. However, the reduction in microbial count in all samples was influenced by the storage temperature, where the lowered microbial count was noted in the sample treated with CE and stored at 5 and 10 °C for 48 h. The pH of meat treated with or without CE ranged from pH 5.74 to 6.48. The sensory attributes of colour, texture, and overall acceptability have a significant difference, except for odour, between marinated meat and control. The results indicate that the use of cinnamon extract as the marination agent for meat could reduce the growth of Pseudomonas spp. and therefore assist in extending the shelf life of meat at 5 and 10 °C storage temperatures.
Fulltext The effects of different packaging materials, temperatures and water activities to control aflatoxin B1 production by Aspergillus flavus and A. parasiticus in stored peanuts

John JM, Jinap S, Hanani ZAN, Nor-Khaizura MAR, Samsudin NIP

J Food Sci Technol, 2019 Jun;56(6):3145-3150.
PMID: 31205369 DOI: 10.1007/s13197-019-03652-6

Aflatoxins (AFs) are secondary metabolites produced by aflatoxigenic strains of Aspergillus flavus and A. parasiticus, the most toxic being aflatoxin B1 (AFB1). The purpose of the present work was to investigate the effects of industrial-grade packaging materials (low-density polyethylene, polypropylene, polyethylene-laminated aluminium); temperatures (25 °C, 30 °C); and water activities (0.74 a w, 0.85 a w) on AFB1 production by A. flavus and A. parasiticus in stored peanut kernels. Commercially-obtained samples were segregated into packaging materials, separately inoculated with the aflatoxigenic Aspergillus spp., and stored for 1 month under various °C + a w regimes. AFB1 production was quantified by high performance liquid chromatography with fluorescence detector (HPLC-FLD). For A. flavus in PELA, no AFB1 was detected (100% reduction) at 25 °C for both a w tested. For A. parasiticus in PELA, no AFB1 was detected at 25 °C (0.85 a w) and 30 °C (0.74 a w). Highest concentration of AFB1 was detected in LDPE for both A. flavus (46.41 ppb) and A. parasiticus (414.42 ppb), followed by PP (A. flavus 24.29 ppb; A. parasiticus 386.73 ppb). In conclusion, storing peanut kernels in PELA in a dry place at room temperature has been demonstrated as an adequate and inexpensive method in inhibiting growth of Aspergillus spp. and lowering AFB1 contamination in peanuts.
Fulltext Aspergillus section Flavi and Aflatoxins: Occurrence, Detection, and Identification in Raw Peanuts and Peanut-Based Products Along the Supply Chain

Norlia M, Jinap S, Nor-Khaizura MAR, Radu S, Samsudin NIP, Azri FA

Front Microbiol, 2019;10:2602.
PMID: 31824445 DOI: 10.3389/fmicb.2019.02602

Aflatoxin contamination in foods is a global concern as they are carcinogenic, teratogenic and mutagenic compounds. The aflatoxin-producing fungi, mainly from the Aspergillus section Flavi, are ubiquitous in nature and readily contaminate various food commodities, thereby affecting human's health. The incidence of aflatoxigenic Aspergillus spp. and aflatoxins in various types of food, especially raw peanuts and peanut-based products along the supply chain has been a concern particularly in countries having tropical and sub-tropical climate, including Malaysia. These climatic conditions naturally support the growth of Aspergillus section Flavi, especially A. flavus, particularly when raw peanuts and peanut-based products are stored under inappropriate conditions. Peanut supply chain generally consists of several major stakeholders which include the producers, collectors, exporters, importers, manufacturers, retailers and finally, the consumers. A thorough examination of the processes along the supply chain reveals that Aspergillus section Flavi and aflatoxins could occur at any step along the chain, from farm to table. Thus, this review aims to give an overview on the prevalence of Aspergillus section Flavi and the occurrence of aflatoxins in raw peanuts and peanut-based products, the impact of aflatoxins on global trade, and aflatoxin management in peanuts with a special focus on peanut supply chain in Malaysia. Furthermore, aflatoxin detection and quantification methods as well as the identification of Aspergillus section Flavi are also reviewed herein. This review could help to shed light to the researchers, peanut stakeholders and consumers on the risk of aflatoxin contamination in peanuts along the supply chain.
Chitosan nanoparticles incorporate with curry leaf essential oil: Physicochemical characterization and in vitro release properties

Nur Fatin Nazurah R, Noranizan MA, Nor-Khaizura MAR, Nur Hanani ZA

Int J Biol Macromol, 2024 Jul;273(Pt 2):132972.
PMID: 38876241 DOI: 10.1016/j.ijbiomac.2024.132972

The use of essential oils as natural antioxidant, antimicrobial and insect repellent agent was limited by the loss of bioactive components especially volatile compounds. This study aimed to improve biological properties of curry leaf essential oil (CLEO) by producing nanometer sized particles through two different synthesis techniques; nanoencapsulation and nanoprecipitation. The methods produced different nanostructures; nanocapsules and nanospheres distinguished by the morphological structure (TEM analysis). Successful loading of CLEO into chitosan nanocarrier was proven by FTIR spectra. Zeta potential values for both nanostructures were more than +30 mV implying their stability against aggregation. CLEO loaded nanocapsules exhibited highest antibacterial properties against Gram-positive bacteria compared to nanospheres. Meanwhile, CLEO loaded nanospheres recorded up until 90.44 % DPPH radical scavenging properties, higher compared to nanocapsules. Both nanostructures demonstrated further improvement in antioxidant and antibacterial activities with the incorporation of higher chitosan concentration. In vitro release analysis indicated that CLEO undergo two-stage discharge mechanism where fast discharge occurred up until 12 h followed by sustained released afterwards. The two synthesis methods applied synergistically with greater chitosan concentration successfully produced nanostructures with >60 % encapsulation efficiency (EE). This concluded that both techniques were reliable to protect the bioactive constituents of CLEO for further used.
Fulltext Assessing the Performance of Food Safety Management System Using Food Safety Management System Diagnostic Tools and Microbial Assessment Scheme: A Case of Powdered Beverage Manufacturers

Cheah HY, Merican SE, Nor Khaizura MAR, Bakar AZA, Omar S, Sanny M

Malays J Med Sci, 2021 Jun;28(3):129-142.
PMID: 34285651 DOI: 10.21315/mjms2021.28.3.12

Background: The objective of the study is to assess the performance of the Food Safety Management System (FSMS) among powdered beverage manufacturers using Food Safety Management System Diagnostic Tools (FSMS-DI) and Microbial Assessment Scheme (MAS).
Methods: FSMS-DI was used to evaluate the context factors, core control and core assurance activities of five powdered beverage manufacturers with different types of FSMS certification. Manufacturer A is not certified with any FSMS, while manufacturers B, C, D and E are complied with MeSTI, GMP, HACCP and ISO 22000, respectively. For MAS, samples were collected from the selected critical sampling locations of two manufacturers who complied FSMS with the least (manufacturer B) and the most stringent (manufacturer E) requirements. The samples consisted of two different types of powdered beverage products were analysed for total plate count (TPC), Salmonella, Escherichia coli, Staphylococcus aureus, yeast and mould count (YMC). Results: The food safety (FS) output of powdered beverages for manufacturer E was better (overall score of 3) than manufacturer B (overall score of 2-3). Manufacturer E was able to achieve their FS objectives. The FSMS activities of manufacturer C, D and E were better (overall score of 2-3) than manufacturer A and B (overall score of 1-2).
Conclusion: The study demonstrated that FSMS-DI and MAS can be used to differentiate the FSMS performance of powdered beverage manufacturers with different types of FSMS certification. Higher scores of FSMS activities obtained by the manufacturer who complied with stringent FSMS certifications contributed to better microbiological safety performance of powdered beverages.
Modelling the effect of temperature and water activity on the growth rate of Aspergillus flavus and aflatoxin production in peanut meal extract agar

Norlia M, Jinap S, Nor-Khaizura MAR, Radu S, John JM, Rahman MAH, et al.

Int J Food Microbiol, 2020 Dec 16;335:108836.
PMID: 33065380 DOI: 10.1016/j.ijfoodmicro.2020.108836

Aspergillus flavus is the predominant species that produce aflatoxins in stored peanuts under favourable conditions. This study aimed to describe the growth and aflatoxin production by two A. flavus strains isolated from imported raw peanuts and to model the effects of temperature and aw on their colony growth rate as a function of temperature and aw in Peanut Meal Extract Agar (PMEA). A full factorial design with seven aw levels (0.85-0.98 aw) and five temperature levels (20-40 °C) was used to investigate the growth and aflatoxin production. Colony diameter was measured daily for 28 days while AFB1 and total aflatoxin were determined on day 3, 7, 14, and 21. The maximum colony growth rate, μmax (mm/day) was estimated by using the primary model of Baranyi, and the μmax was then fitted to the secondary model; second-order polynomial and linear Arrhenius-Davey to describe the colony growth rate as a function of temperature and aw. The results indicated that both strains failed to grow at temperature of 20 °C with aw <0.94 and aw of 0.85 for all temperatures except 30 °C. The highest growth rate was observed at 30 °C, with 0.98 aw for both strains. The analysis of variance showed a significant effect of strain, temperature, and aw on the fungal growth and aflatoxin production (p
Characterization, molecular identification, and antimicrobial activity of lactic acid bacteria isolated from selected fermented foods and beverages in Malaysia

Haryani Y, Halid NA, Guat GS, Nor-Khaizura MAR, Hatta A, Sabri S, et al.

FEMS Microbiol Lett, 2023 Jan 17;370.
PMID: 37002414 DOI: 10.1093/femsle/fnad023

The present work investigated the profile and biodiversity of lactic acid bacteria (LAB) isolated from selected manufactured and homemade fermented foods in Malaysia. A total of 55 LAB were isolated from 20 samples, and identified based on the sequencing of 16S rRNA gene. The LAB isolates were identified as Lacticaseibacillus rhamnosus (34.5%), Lactiplantibacillus plantarum (20%), Limosilactobacillus fermentum (20%), Lacticaseibacillus paracasei (12.7%), Lacticaseibacillus casei (3.6%), Lactobacillus sp. (1.8%), Enterococcus faecalis (3.6%), Enterococcus faecium (1.8%), and Enterococcus durans (1.8%). Majority (94%) of the LAB isolates exhibited broad-spectrum antimicrobial activity against selected foodborne pathogens, and four isolates (L. fermentum SC1001, L. paracasei K2003, and L. rhamnosus KF1002 and MK2003) could produce bacteriocin-like inhibitory substance (BLIS). Lacticaseibacillus paracasei M1001 (homemade mozzarella) exhibited high-temperature tolerance and acid resistance, was homofermentative, and generated good antimicrobial activity, which strongly implied its potential for industrial applications. The present work results would potentially widen our knowledge of LAB diversity in Malaysian fermented foods and provide a potential for their applications in the food industry or other purposes.
Efficient Metabolic Pathway Modification in Various Strains of Lactic Acid Bacteria using CRISPR/Cas9 System for elevated synthesis of Antimicrobial Compounds

Haryani Y, Halid NA, Guat GS, Nor-Khaizura MAR, Hatta MAM, Sabri S, et al.

J Biotechnol, 2024 Sep 06.
PMID: 39245212 DOI: 10.1016/j.jbiotec.2024.09.002

Lactic acid bacteria (LAB) are known to exhibit various beneficial roles in fermentation, serving as probiotics, and producing a plethora of valuable compounds including compounds with antimicrobial activity including bacteriocin-like inhibitory substance (BLIS) that can be used as biopreservative to improve food safety and quality. However, the yield of BLIS is often limited, which poses a challenge to be commercially competitive with the current preservation practice. Therefore, the present work aimed to establish an optimised two-plasmid CRISPR/Cas9 system to redirect the carbon flux away from lactate towards compounds with antimicrobial activity by disrupting lactate dehydrogenase gene (ldh) on various strains of LAB. The lactic acid-deficient (ldhΔ) strains caused a metabolic shift resulting in increased inhibitory activity against selected foodborne pathogens up to 78% than the wild-type (WT) strain. The most significant effect was depicted by Enterococcus faecalis-ldh∆ which displayed prominent bactericidal effects against all foodborne pathogens as compared to the WT that showed no antimicrobial activity. The present work provided a framework model for economically important LAB and other beneficial bacteria to synthesise and increase the yield of valuable food and industrial compounds. The present work reported for the first time that the metabolism of selected LAB can be manipulated by modifying ldh to attain metabolites with higher antimicrobial activity.
Formaldehyde contamination in seafood industry: an update on detection methods and legislations

Mehta NK, Vaishnav A, Priyadarshini MB, Debbarma P, Hoque MS, Mondal P, et al.

Environ Sci Pollut Res Int, 2024 Sep 03.
PMID: 39223414 DOI: 10.1007/s11356-024-34792-8

Seafood is abundant in high-quality protein, healthy fats (n-3 and n-6 PUFAs), minerals (calcium, magnesium, copper, selenium, and so on), and vitamin D. Functional compounds in seafood can protect against lifestyle-related diseases. Having had all the merits mentioned, it is also a highly putrefiable food item. To maintain quality and prolong seafood's shelf life, various chemicals have been added, including nitrite, sulfur dioxide, and formaldehyde. In this review, we summarize the uses, the incidence of added formalin contamination, and the approved limit for seafood products. Additionally, worldwide regulations or standards for the use of formalin in seafood products, as well as recent changes relevant to new methods, are highlighted. Although strict limits and regulations have been placed on the utilization of formaldehyde for seafood preservation, there are few incidences reported of formalin/formaldehyde detection in seafood products around Asian countries. In this context, various qualitative and quantitative detection methods for formaldehyde have been developed to ensure the presence of formaldehyde within acceptable limits. Besides this, different rules and regulations have been forced by each country to control formaldehyde incidence. Although it is not an issue of formaldehyde incidence in European countries, strict regulations are implemented and followed.

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