# The Occurrence of Aflatoxins in Date Palm (Phoenix dactylifera L.) Worldwide

Authors: Merfat Abdulrahman Almaghrabi
Journal: Journal of Food Quality
 (2022)

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## Abstract

Due to global warming, the risk of aflatoxins exposure through the consumption of contaminated food has increased. Aflatoxins pose serious health hazards to humans’ and animals’ health because of their carcinogenic, mutagenic, and teratogenic properties and their immunosuppressive effects. Aflatoxin contamination in various agricultural commodities has attracted much attention worldwide. Date palm fruits are among these important commodities that are vulnerable to fungal contamination and consequent aflatoxins production. Furthermore, dates are often consumed directly without any further processing, which may result in direct exposure to aflatoxins. Moreover, dates are the second dried fruits traded worldwide, which reflects the widespread consumption of dates due to their nutritive values in addition to religious and cultural values. Accordingly, this review summarizes and discusses the frequency and incidence of aflatoxin contamination in dates worldwide and outlines the analytical procedure for aflatoxin determination in dates for the first time. The susceptibility of date palm fruits to aflatoxins contamination has been documented at various levels in several regions. The findings urged the importance of conducting more comprehensive studies on aflatoxin occurrence and contamination levels in dates as a likely contributor to the dietary exposure to aflatoxins.


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## Body

## 1. Introduction

Food security and the potential effects of global warming on crops have become crucial aspects worldwide [1]. Global warming has increased the risk of mycotoxin exposure as the Earth’s temperature has increased; water evaporation from the Earth’s surface has increased as well, resulting in escalating humidity and spreading the risk of mycotoxigenic Aspergillus species. Consequently, these climatic changes have increased the production of mycotoxins, such as aflatoxins, which are among the most hazardous mycotoxins [2, 3].More than 20 forms of aflatoxins exist. However, aflatoxin B1 (AFB1), aflatoxin B2 (AFB2), aflatoxin G1 (AFG1), and aflatoxin G2 (AFG2) are considered the most common and important types of aflatoxins [4]. These toxins contaminate various agricultural commodities, such as nuts, spices, cereals, and dried fruits [5, 6]. A recent risk assessment of aflatoxins in food published by the European Food Safety Authority (EFSA), which assessed the toxicity of aflatoxins and subsequently their health risks to humans, reported that, among all types of aflatoxins, AFB1 was the most frequent contaminant detected in food, and it is not appropriate to establish a tolerable daily intake (TDI) for aflatoxins [7]. This is because aflatoxins pose a risk to human and animal health due to their carcinogenic, mutagenic, and teratogenic properties and immunosuppressive effects [7–9]. The consequences of low-dose exposure over a long period of time or high-dose exposure over a short time include chronic toxicity or acute toxicity and death, respectively, in humans [10]. The primary target organ of aflatoxin is the liver, and aflatoxin can be metabolised by the cytochrome P450 (CYP) enzyme system. The CYP enzymes convert AFB1, AFG1, and AFM1 to their respective epoxides, which can bind covalently to both DNA and proteins. In particular, aflatoxin epoxide can bind rapidly to the N-7 position of guanine in DNA to form AFB1-N7-guanine adducts, which are mutagenic if not repaired, while aflatoxin epoxide in liver cells and aflatoxin dialdehyde in blood can bind covalently to lysine in albumin to form aflatoxin albumin adducts [11]. The International Agency for Research on Cancer (IARC) lists AFB1, AFB2, AFG1, AFG2, and AFM1 as Group 1 carcinogens based on strong evidence of their genotoxicity being involved in the formation of DNA adducts causing point mutations in the TP53 gene [12]. The EFSA emphasises the importance of continuously monitoring the occurrence of aflatoxins due to the Earth’s changing climate and its impact on increases in the risk of aflatoxins contamination [7].The presence of aflatoxins requires warm temperatures and high humidity [13], such as in tropical, subtropical, and Mediterranean climates [14, 15]. The date palm (Phoenix dactylifera L.) is a crop grown in these regions [16–18], and it is also cultivated in its preferred habitats in the Middle East, North Africa, and the Arabian Peninsula [19]. According to the database of the Food and Agriculture Organization of the United Nations [20], the lead date-producing countries over the period of 2010 to 2019 were Egypt, Iran, Saudi Arabia, Algeria, Iraq, Pakistan, Sudan, United Arab Emirates, Oman, and Tunisia (Figure S1). Despite the importance of date fruits in these regions and also their significance for Muslims, particularly during the holy month of Ramadan [21], the consumption of dates has increased outside of production areas [22]. As a result, dates are the second dried fruits traded throughout the world after raisins due to their sweet taste and nutritive values [22]. Figures 1 and 2 reflect the demand for dates worldwide and the widespread consumption of dates based on data extracted from the International Trade Centre over the period of 2010 to 2019 [23].Figure 1
Dates global total import in tonnes for the period of 2010–2019. The data extracted from the International Trade Centre database [23].Figure 2
Dates global average import in tonnes by the highest importing countries of dates for the period of 2010–2019. The data extracted from the International Trade Centre database [23].Dates are rich in carbohydrates, some amino acids, and fiber, and they also contain significant amounts of vitamins and various minerals, particularly iron, potassium, calcium, and magnesium. Consequently, dates are among the most nourishing natural foods in the human diet [24]. However, dates also provide an ideal medium for toxigenic fungi growth and further aflatoxins contamination [16, 17, 22, 25]. Furthermore, dates are grown in climatic conditions that facilitate the toxigenic fungal invasion and aflatoxin formation [16, 17, 22, 26].Such fungal invasion and the incidence of aflatoxins in date fruits have been investigated and confirmed in several countries, but with less attention compared with numerous publications on aflatoxins contamination in other agricultural commodities. The Food and Agriculture Organization (FAO) estimated the worldwide loss of dates in 2019 to be 1000 tonnes, excluding the losses that occurred before and during harvest [27]. Suhail et al. [28] investigated the postharvest losses of dates along the supply chain in Pakistan and found that the total losses of dates at the farm level were 29.6%, compared with 41.33% and 35.33% at wholesale and retail levels, respectively. The results demonstrated that fungal attacks are one of the main reasons for date loss. Atia [29] reported that the most common fungi attributed to the loss of the dates is the Aspergillus species. Until now, the available data on the losses of this valuable food stuffs due to aflatoxin contamination has been limited.Monitoring the occurrence of aflatoxins contamination in dates is crucial, as dates are often consumed directly without any further processing, which may result in direct exposure to aflatoxins and pose human health problems. Therefore, the purpose of the current review is to summarize the available data regarding aflatoxin occurrence and contamination levels in date fruits worldwide and to outline the analytical procedures used for aflatoxin determination in dates.


## 2. Methodology

### 2.1. Criteria Used to Select Articles

An electronic search was conducted in major databases including Web of Science, PubMed, and Google Scholar using “Aflatoxin∗ AND Dried fruits”, “Aflatoxin∗ AND Date fruits”, Aflatoxin∗ AND Date Palm and “Mycotoxin∗ AND Dried fruits” as keywords. The databases were screened for the period between 2010 and 2019 with English language. The electronic search identified 170 articles, of which 14 were found to be eligible for inclusion. The publications were evaluated through the title, abstract, and full text of papers to ensure that these studies covered levels of aflatoxins contamination in dates (Figure 3).Figure 3
Flowchart indicating the search and selection process of articles.


## 2.1. Criteria Used to Select Articles

An electronic search was conducted in major databases including Web of Science, PubMed, and Google Scholar using “Aflatoxin∗ AND Dried fruits”, “Aflatoxin∗ AND Date fruits”, Aflatoxin∗ AND Date Palm and “Mycotoxin∗ AND Dried fruits” as keywords. The databases were screened for the period between 2010 and 2019 with English language. The electronic search identified 170 articles, of which 14 were found to be eligible for inclusion. The publications were evaluated through the title, abstract, and full text of papers to ensure that these studies covered levels of aflatoxins contamination in dates (Figure 3).Figure 3
Flowchart indicating the search and selection process of articles.


## 3. Results

### 3.1. Analytical Procedure for Aflatoxins Determination in Date Palm Fruits

The general trend in aflatoxins analysis is the performance of total aflatoxins (AFB1, AFB2, AFG1, and AFG2) methods, which allow the analysis of a wide range of aflatoxins in a single run, using liquid chromatography coupled to mass spectrometry/mass (LC-MS/MS) technique [30–34]. In addition, high performance liquid chromatography (HPLC) equipped with fluorescence detection was used in six studies [21, 26, 35–38]. These studies used an immunoaffinity column (IAC) for clean-up. The Kobra cell and trifluoroacetic acid (TFA) were used as derivatisation to increase the detection sensitivity (Table 1). In the study of Hegazy and El Sayed [40], enzyme-linked immunosorbent assay (ELISA) method was used for quantified aflatoxins levels using aflatoxin test kits. Based on this review, acetonitrile followed by methanol were the main elements of the polar aqueous media used in the extraction of aflatoxins from date palm fruits (Table 1). The adequacy of different analytical procedures utilised to determine aflatoxins in date palm fruits in the current review was indicated by the method’s sensitivity. Particularly, the determined limits of detections (LODs) of the method, apart from the studies of Ibrahim et al. [35] and Azaiez et al. [31, 34], were not reported. The LODs of the analytical procedure are presented in Table 1. The reported LODs for aflatoxins ranged from 1 to 0.04 μg/kg, 0.03 μg/kg, 0.04 μg/kg, and 0.02 μg/kg for AFB1, AFB2, AFG1, and AFG2, respectively. These values are below all current aflatoxins legislation, thus revealing all determined aflatoxin levels that exceed the permissible limits of aflatoxin specified in different legislation (Table 2).Table 1
Analytical procedure for aflatoxins determination in date palm fruits.
CountryTechDerivatizationExtractionClean-upLimit of detection µg/kgReferenceB1B2G1G2Pakistan (Khyber Pakhtunkhwa)LCTFAMethanol: waterImmunoaffinity column0.51.00.51.0[39]Saudi Arabia (Riyadh)HPLCPBPBMethanol: waterImmunoaffinity column————[35]Pakistan (Punjab and Khyber Pakhtunkhwa)HPLCTFAAcetonitrile: waterImmunoaffinity column0.050.060.050.06[21]Egypt (Cairo)ELISA—Methanol: water—1.0[40]Spain (Valencia)LC-MS/MS—Acetonitrile: water: acetic acid—0.080.080.160.3[30]Pakistan (Khyber Pakhtunkhwa)HPLCTFAAcetonitrile: waterImmunoaffinity column0.040.070.040.07[36]Spain (Valencia)LC-MS/MS—Acetonitrile: water: acetic acid—————[31]Tunisia (Tunis)LC-MS/MS—Acetonitrile: water: acetic acid—————[31]China (Shanghai)LC-MS/MS—Acetonitrile: water—0.10.10.30.3[32]Pakistan (different areas of Pakistan)HPLCKobra Cell™Methanol: waterImmunoaffinity column0.12[26]Iran (Hamadan)HPLC—Methanol: waterImmunoaffinity column0.060.050.080.02[37]Pakistan (Punjab and Khyber Pakhtunkhwa)HPLCTFAAcetonitrile: waterImmunoaffinity column0.040.070.040.07[38]Egypt (Assiut)LC-MS/MS—Acetonitrile: water: acetic acid—0.050.03——[33]Italy (Perugia)LC-MS/MS—Acetonitrile: water: acetic acid—————[34]LC-MS/MS: liquid chromatography with tandem mass spectrometry; HPLC: high performance liquid chromatography; ELISA: enzyme-linked immunosorbent assay; TFA: trifluoroacetic acid; PBPB: pyridinium hydrobromide perbromide.Table 2
Legal limits for aflatoxins of dried fruit in gulf standardization organization, European Union member states, and the United States, compared with the studies reporting the occurrence of aflatoxins in date palm fruits.
AflatoxinsMaximum acceptable levels in Gulf Standardization Organization (μg/kg)Maximum acceptable levels in EU member states (μg/kg)Maximum acceptable levels in the United States (μg/kg)(GSO)(EU)(FDA)[41][42][43]AFB1——2∗5∗∗—Total aflatoxins (AFB1, AFB2, AFG1, and AFG2)4∗10∗∗4∗10∗∗20Studies reported average AFB1 and AFs (μg/kg) compared with maximum legal limits (MLL)Pakistan[39]AFs (2.5 < 4–10, GSO and EU MLL)AFs (2.5 < 20, FDA MLL)Saudi Arabia[35]AFB1 (1.39 < 2–5, EU MLL)AFs (2.01 < 20, FDA MLL)AFs (2.01 < 4–10, GSO and EU MLL)Pakistan[21]AFB1 (2.13 ≥ 2–5, EU MLL)AFs (4.11 < 20, FDA MLL)AFs (4.11 ≥ 4–10, GSO and EU MLL)Egypt[40]——Spain[30]——Pakistan[36]AFB1 (4.50 > 2, EU MLL)AFs (6.32 < 20, FDA MLL)AFB1 (4.50 < 5, EU MLL)AFs (6.32 > 4, GSO and EU MLL)AFs (6.32 < 10, GSO and EU MLL)Spain[31]——Tunisia[31]AFs (4.24 ≥ 4–10, GSO and EU MLL)AFs (4.24 < 20, FDA MLL)China[32]——Pakistan[26]AFs (0.24 < 4–10, GSO and EU MLL)AFs (0.24 < 20, FDA MLL)Iran[37]AFB1 (2.1 ≤ 2–5, EU MLL)AFs (2.6 < 20, FDA MLL)AFs (2.6 < 4–10, GSO and EU MLL)Pakistan[38]AFB1 (3.20 and 4.80 > 2, EU MLL)AFs (3.90 and 5.30 < 20, FDA MLL)AFB1 (3.20 and 4.80 < 5, EU MLL)AFs (3.90 < 4–10, GSO and EU MLL)AFs (5.30 > 4, GSO and EU MLL)AFs(5.30 < 10, GSO and EU MLL)Egypt[33]AFB1 (14.4 > 2–5, EU MLL)AFs (16.84 < 20, FDA MLL)AFs (16.84 > 4–10, GSO and EU MLL)Italy[34]——∗Dried fruit and processed products thereof, intended for direct human consumption or use as an ingredient in foodstuff. ∗∗Dried fruit to be subjected to sorting or other physical treatment before human consumption or use as an ingredient in foodstuffs.


### 3.2. Levels and Incidence

The frequency and the incidence of aflatoxins contamination in dates in various world regions are presented in Table3.Table 3
The frequency and the incidence of aflatoxins contamination in dates in various world regions.
CountryNo. of samples/contaminated samplesAflatoxinsAverage (µg/kg)Range of contamination (µg/kg)ReferencePakistan (Khyber Pakhtunkhwa)20/2 (dried dates)Total AFs2.52.1–2.9[39]Saudi Arabia (Riyadh)12/10 (Semi-dry dates)AFB11.39 ± 1.420.00 to 5.92[35]Total AFs2.01 ± 1.76Pakistan (Punjab and Khyber Pakhtunkhwa)96/38 (different date varieties)AFB12.13 ± 0.62LOD-26.60[21]Total AFs4.11 ± 0.80Egypt (Cairo)5/0 (rutab and sukkary)Total AFsAbsAbs[40]Spain (Valencia)3/0 (dried dates)Total AFsAbsAbs[30]Pakistan (Khyber Pakhtunkhwa)15/9 (dried dates)AFB14.50 ± 0.46LOD-9.80[36]Total AFs6.32 ± 0.78LOD-18.79Spain (Valencia)27/0 (different date varieties)Total AFsAbsAbs[31]Tunisia (Tunis)48/22 (different date varieties)AFB1n.d.Abs[31]AFB21.141.1–1.3AFG11.4LOQ-1.8AFG21.7LOQ- 2.2China (Shanghai)40/0 (dried dates)Total AFsAbsAbs[32]Pakistan (different areas of Pakistan)170/25 (dried dates)Total AFs0.24 ± 0.080.24–5.87[26]Iran (Hamadan)22/9 (dried dates)AFB12.1 ± 1.80.6–6[37]Total AFs2.6 ± 2.30.9–8.1Pakistan (Punjab and Khyber Pakhtunkhwa)15/6 (dates)AFB14.80 ± 0.95LOD-10.20[38]Total AFs5.30 ± 1.8017/5 (dried dates)AFB13.20 ± 0.90LOD-15.50Total AFs3.90 ± 0.95Egypt (Assiut)28/1 (dried dates)AFB1Abs14.4[33]AFB2Abs2.44Italy (Perugia)20/0 (dried dates)Total AFsAbsAbs[34]LOD: limit of detection. LOQ: limit of quantification. n.d.: not detectable. Abs: absent.


#### 3.2.1. Asia

A number of studies have reported the occurrence of aflatoxins in dates in various regions in Pakistan. Iqbal et al. reported the incidence of AFB1 and total aflatoxins (AFs) in Punjab and Khyber Pakhtunkhwa in 2014 and 2018. The finding of 2014 study demonstrated that aflatoxins were detected in 39.6% of different date varieties, with an average of 2.13 μg/kg and 4.11 μg/kg for AFB1 and total AFs, respectively [21]. However, in 2018, a lower level of total AFs was found in dates with an average of 3.90 μg/kg [38]. Another investigation conducted in Khyber Pakhtunkhwa, Pakistan, detected aflatoxins in only 10% of samples, with an average of 2.5 μg/kg [39]. However, Masood et al. [36] have reported very high incidence rates in Khyber Pakhtunkhwa; that is, 60% date samples were contaminated with an average of 4.50 μg/kg and 6.32 μg/kg for AFB1 and total Afs, respectively. Over the period under review, the highest documented survey concerning aflatoxins contamination of dates was conducted in Pakistan from 2012 to 2015. Out of 170 collected samples, 25 were positive, with an average of 0.24 μg/kg for total Afs [26].Aflatoxin contamination levels in Iranian dates were similar to the reported incidence in dates marketed in Riyadh, Saudi Arabia, where the mean concentration is about 2μg/kg [35, 37], although higher frequency was documented in Saudi Arabia with 80% contamination. In Shanghai, China, none of Afs types were detected in all 40 samples of dates investigated by Han et al. [32].


#### 3.2.2. Europe

In Europe, there were no aflatoxins detected in date samples collected from Valencia, Spain, and Perugia, Italy [30, 34]. Additionally, the findings from the study of Azaiez et al. [31] reported that there was no incidence of aflatoxins contamination in various date varieties marketed in Spain. These samples were imported from various countries, namely, Tunisia, Israel, and Algeria.


#### 3.2.3. Africa

Limited reports have been published regarding the incidence of aflatoxins contamination in dates in Africa. Hegazy and El Sayed [40] from Egypt investigated the presence of aflatoxins in dates (rutab and sukkary) and found that the samples were free from aflatoxins. The results are consistent with recent findings from Abdallah et al. [33], who found only one sample contaminated at levels of 14.4 μg/kg and 2.44 μg/kg for AFB1 and AFB2, respectively. In Tunisian markets, several date varieties (48 samples) were collected and analyzed; the average contamination levels of AFs in 22 positive samples were 1.14 μg/kg, 1.4 μg/kg, and 1.7 μg/kg for AFB2, AFG1, and AFG2, respectively [31].


## 3.1. Analytical Procedure for Aflatoxins Determination in Date Palm Fruits

The general trend in aflatoxins analysis is the performance of total aflatoxins (AFB1, AFB2, AFG1, and AFG2) methods, which allow the analysis of a wide range of aflatoxins in a single run, using liquid chromatography coupled to mass spectrometry/mass (LC-MS/MS) technique [30–34]. In addition, high performance liquid chromatography (HPLC) equipped with fluorescence detection was used in six studies [21, 26, 35–38]. These studies used an immunoaffinity column (IAC) for clean-up. The Kobra cell and trifluoroacetic acid (TFA) were used as derivatisation to increase the detection sensitivity (Table 1). In the study of Hegazy and El Sayed [40], enzyme-linked immunosorbent assay (ELISA) method was used for quantified aflatoxins levels using aflatoxin test kits. Based on this review, acetonitrile followed by methanol were the main elements of the polar aqueous media used in the extraction of aflatoxins from date palm fruits (Table 1). The adequacy of different analytical procedures utilised to determine aflatoxins in date palm fruits in the current review was indicated by the method’s sensitivity. Particularly, the determined limits of detections (LODs) of the method, apart from the studies of Ibrahim et al. [35] and Azaiez et al. [31, 34], were not reported. The LODs of the analytical procedure are presented in Table 1. The reported LODs for aflatoxins ranged from 1 to 0.04 μg/kg, 0.03 μg/kg, 0.04 μg/kg, and 0.02 μg/kg for AFB1, AFB2, AFG1, and AFG2, respectively. These values are below all current aflatoxins legislation, thus revealing all determined aflatoxin levels that exceed the permissible limits of aflatoxin specified in different legislation (Table 2).Table 1
Analytical procedure for aflatoxins determination in date palm fruits.
CountryTechDerivatizationExtractionClean-upLimit of detection µg/kgReferenceB1B2G1G2Pakistan (Khyber Pakhtunkhwa)LCTFAMethanol: waterImmunoaffinity column0.51.00.51.0[39]Saudi Arabia (Riyadh)HPLCPBPBMethanol: waterImmunoaffinity column————[35]Pakistan (Punjab and Khyber Pakhtunkhwa)HPLCTFAAcetonitrile: waterImmunoaffinity column0.050.060.050.06[21]Egypt (Cairo)ELISA—Methanol: water—1.0[40]Spain (Valencia)LC-MS/MS—Acetonitrile: water: acetic acid—0.080.080.160.3[30]Pakistan (Khyber Pakhtunkhwa)HPLCTFAAcetonitrile: waterImmunoaffinity column0.040.070.040.07[36]Spain (Valencia)LC-MS/MS—Acetonitrile: water: acetic acid—————[31]Tunisia (Tunis)LC-MS/MS—Acetonitrile: water: acetic acid—————[31]China (Shanghai)LC-MS/MS—Acetonitrile: water—0.10.10.30.3[32]Pakistan (different areas of Pakistan)HPLCKobra Cell™Methanol: waterImmunoaffinity column0.12[26]Iran (Hamadan)HPLC—Methanol: waterImmunoaffinity column0.060.050.080.02[37]Pakistan (Punjab and Khyber Pakhtunkhwa)HPLCTFAAcetonitrile: waterImmunoaffinity column0.040.070.040.07[38]Egypt (Assiut)LC-MS/MS—Acetonitrile: water: acetic acid—0.050.03——[33]Italy (Perugia)LC-MS/MS—Acetonitrile: water: acetic acid—————[34]LC-MS/MS: liquid chromatography with tandem mass spectrometry; HPLC: high performance liquid chromatography; ELISA: enzyme-linked immunosorbent assay; TFA: trifluoroacetic acid; PBPB: pyridinium hydrobromide perbromide.Table 2
Legal limits for aflatoxins of dried fruit in gulf standardization organization, European Union member states, and the United States, compared with the studies reporting the occurrence of aflatoxins in date palm fruits.
AflatoxinsMaximum acceptable levels in Gulf Standardization Organization (μg/kg)Maximum acceptable levels in EU member states (μg/kg)Maximum acceptable levels in the United States (μg/kg)(GSO)(EU)(FDA)[41][42][43]AFB1——2∗5∗∗—Total aflatoxins (AFB1, AFB2, AFG1, and AFG2)4∗10∗∗4∗10∗∗20Studies reported average AFB1 and AFs (μg/kg) compared with maximum legal limits (MLL)Pakistan[39]AFs (2.5 < 4–10, GSO and EU MLL)AFs (2.5 < 20, FDA MLL)Saudi Arabia[35]AFB1 (1.39 < 2–5, EU MLL)AFs (2.01 < 20, FDA MLL)AFs (2.01 < 4–10, GSO and EU MLL)Pakistan[21]AFB1 (2.13 ≥ 2–5, EU MLL)AFs (4.11 < 20, FDA MLL)AFs (4.11 ≥ 4–10, GSO and EU MLL)Egypt[40]——Spain[30]——Pakistan[36]AFB1 (4.50 > 2, EU MLL)AFs (6.32 < 20, FDA MLL)AFB1 (4.50 < 5, EU MLL)AFs (6.32 > 4, GSO and EU MLL)AFs (6.32 < 10, GSO and EU MLL)Spain[31]——Tunisia[31]AFs (4.24 ≥ 4–10, GSO and EU MLL)AFs (4.24 < 20, FDA MLL)China[32]——Pakistan[26]AFs (0.24 < 4–10, GSO and EU MLL)AFs (0.24 < 20, FDA MLL)Iran[37]AFB1 (2.1 ≤ 2–5, EU MLL)AFs (2.6 < 20, FDA MLL)AFs (2.6 < 4–10, GSO and EU MLL)Pakistan[38]AFB1 (3.20 and 4.80 > 2, EU MLL)AFs (3.90 and 5.30 < 20, FDA MLL)AFB1 (3.20 and 4.80 < 5, EU MLL)AFs (3.90 < 4–10, GSO and EU MLL)AFs (5.30 > 4, GSO and EU MLL)AFs(5.30 < 10, GSO and EU MLL)Egypt[33]AFB1 (14.4 > 2–5, EU MLL)AFs (16.84 < 20, FDA MLL)AFs (16.84 > 4–10, GSO and EU MLL)Italy[34]——∗Dried fruit and processed products thereof, intended for direct human consumption or use as an ingredient in foodstuff. ∗∗Dried fruit to be subjected to sorting or other physical treatment before human consumption or use as an ingredient in foodstuffs.


## 3.2. Levels and Incidence

The frequency and the incidence of aflatoxins contamination in dates in various world regions are presented in Table3.Table 3
The frequency and the incidence of aflatoxins contamination in dates in various world regions.
CountryNo. of samples/contaminated samplesAflatoxinsAverage (µg/kg)Range of contamination (µg/kg)ReferencePakistan (Khyber Pakhtunkhwa)20/2 (dried dates)Total AFs2.52.1–2.9[39]Saudi Arabia (Riyadh)12/10 (Semi-dry dates)AFB11.39 ± 1.420.00 to 5.92[35]Total AFs2.01 ± 1.76Pakistan (Punjab and Khyber Pakhtunkhwa)96/38 (different date varieties)AFB12.13 ± 0.62LOD-26.60[21]Total AFs4.11 ± 0.80Egypt (Cairo)5/0 (rutab and sukkary)Total AFsAbsAbs[40]Spain (Valencia)3/0 (dried dates)Total AFsAbsAbs[30]Pakistan (Khyber Pakhtunkhwa)15/9 (dried dates)AFB14.50 ± 0.46LOD-9.80[36]Total AFs6.32 ± 0.78LOD-18.79Spain (Valencia)27/0 (different date varieties)Total AFsAbsAbs[31]Tunisia (Tunis)48/22 (different date varieties)AFB1n.d.Abs[31]AFB21.141.1–1.3AFG11.4LOQ-1.8AFG21.7LOQ- 2.2China (Shanghai)40/0 (dried dates)Total AFsAbsAbs[32]Pakistan (different areas of Pakistan)170/25 (dried dates)Total AFs0.24 ± 0.080.24–5.87[26]Iran (Hamadan)22/9 (dried dates)AFB12.1 ± 1.80.6–6[37]Total AFs2.6 ± 2.30.9–8.1Pakistan (Punjab and Khyber Pakhtunkhwa)15/6 (dates)AFB14.80 ± 0.95LOD-10.20[38]Total AFs5.30 ± 1.8017/5 (dried dates)AFB13.20 ± 0.90LOD-15.50Total AFs3.90 ± 0.95Egypt (Assiut)28/1 (dried dates)AFB1Abs14.4[33]AFB2Abs2.44Italy (Perugia)20/0 (dried dates)Total AFsAbsAbs[34]LOD: limit of detection. LOQ: limit of quantification. n.d.: not detectable. Abs: absent.


### 3.2.1. Asia

A number of studies have reported the occurrence of aflatoxins in dates in various regions in Pakistan. Iqbal et al. reported the incidence of AFB1 and total aflatoxins (AFs) in Punjab and Khyber Pakhtunkhwa in 2014 and 2018. The finding of 2014 study demonstrated that aflatoxins were detected in 39.6% of different date varieties, with an average of 2.13 μg/kg and 4.11 μg/kg for AFB1 and total AFs, respectively [21]. However, in 2018, a lower level of total AFs was found in dates with an average of 3.90 μg/kg [38]. Another investigation conducted in Khyber Pakhtunkhwa, Pakistan, detected aflatoxins in only 10% of samples, with an average of 2.5 μg/kg [39]. However, Masood et al. [36] have reported very high incidence rates in Khyber Pakhtunkhwa; that is, 60% date samples were contaminated with an average of 4.50 μg/kg and 6.32 μg/kg for AFB1 and total Afs, respectively. Over the period under review, the highest documented survey concerning aflatoxins contamination of dates was conducted in Pakistan from 2012 to 2015. Out of 170 collected samples, 25 were positive, with an average of 0.24 μg/kg for total Afs [26].Aflatoxin contamination levels in Iranian dates were similar to the reported incidence in dates marketed in Riyadh, Saudi Arabia, where the mean concentration is about 2μg/kg [35, 37], although higher frequency was documented in Saudi Arabia with 80% contamination. In Shanghai, China, none of Afs types were detected in all 40 samples of dates investigated by Han et al. [32].


### 3.2.2. Europe

In Europe, there were no aflatoxins detected in date samples collected from Valencia, Spain, and Perugia, Italy [30, 34]. Additionally, the findings from the study of Azaiez et al. [31] reported that there was no incidence of aflatoxins contamination in various date varieties marketed in Spain. These samples were imported from various countries, namely, Tunisia, Israel, and Algeria.


### 3.2.3. Africa

Limited reports have been published regarding the incidence of aflatoxins contamination in dates in Africa. Hegazy and El Sayed [40] from Egypt investigated the presence of aflatoxins in dates (rutab and sukkary) and found that the samples were free from aflatoxins. The results are consistent with recent findings from Abdallah et al. [33], who found only one sample contaminated at levels of 14.4 μg/kg and 2.44 μg/kg for AFB1 and AFB2, respectively. In Tunisian markets, several date varieties (48 samples) were collected and analyzed; the average contamination levels of AFs in 22 positive samples were 1.14 μg/kg, 1.4 μg/kg, and 1.7 μg/kg for AFB2, AFG1, and AFG2, respectively [31].


## 3.2.1. Asia

A number of studies have reported the occurrence of aflatoxins in dates in various regions in Pakistan. Iqbal et al. reported the incidence of AFB1 and total aflatoxins (AFs) in Punjab and Khyber Pakhtunkhwa in 2014 and 2018. The finding of 2014 study demonstrated that aflatoxins were detected in 39.6% of different date varieties, with an average of 2.13 μg/kg and 4.11 μg/kg for AFB1 and total AFs, respectively [21]. However, in 2018, a lower level of total AFs was found in dates with an average of 3.90 μg/kg [38]. Another investigation conducted in Khyber Pakhtunkhwa, Pakistan, detected aflatoxins in only 10% of samples, with an average of 2.5 μg/kg [39]. However, Masood et al. [36] have reported very high incidence rates in Khyber Pakhtunkhwa; that is, 60% date samples were contaminated with an average of 4.50 μg/kg and 6.32 μg/kg for AFB1 and total Afs, respectively. Over the period under review, the highest documented survey concerning aflatoxins contamination of dates was conducted in Pakistan from 2012 to 2015. Out of 170 collected samples, 25 were positive, with an average of 0.24 μg/kg for total Afs [26].Aflatoxin contamination levels in Iranian dates were similar to the reported incidence in dates marketed in Riyadh, Saudi Arabia, where the mean concentration is about 2μg/kg [35, 37], although higher frequency was documented in Saudi Arabia with 80% contamination. In Shanghai, China, none of Afs types were detected in all 40 samples of dates investigated by Han et al. [32].


## 3.2.2. Europe

In Europe, there were no aflatoxins detected in date samples collected from Valencia, Spain, and Perugia, Italy [30, 34]. Additionally, the findings from the study of Azaiez et al. [31] reported that there was no incidence of aflatoxins contamination in various date varieties marketed in Spain. These samples were imported from various countries, namely, Tunisia, Israel, and Algeria.


## 3.2.3. Africa

Limited reports have been published regarding the incidence of aflatoxins contamination in dates in Africa. Hegazy and El Sayed [40] from Egypt investigated the presence of aflatoxins in dates (rutab and sukkary) and found that the samples were free from aflatoxins. The results are consistent with recent findings from Abdallah et al. [33], who found only one sample contaminated at levels of 14.4 μg/kg and 2.44 μg/kg for AFB1 and AFB2, respectively. In Tunisian markets, several date varieties (48 samples) were collected and analyzed; the average contamination levels of AFs in 22 positive samples were 1.14 μg/kg, 1.4 μg/kg, and 1.7 μg/kg for AFB2, AFG1, and AFG2, respectively [31].


## 4. Legal Limit

In regulation terms, there is no harmonized regulation for aflatoxins on a global level [44, 45]. However, aflatoxins are among the most important contaminates regulated by several countries and organizations to reduce the risk posed by these compounds to human health [46]. Table 2 presents a summary of the legal limits for aflatoxins in dried fruit according to the Gulf Standardization Organization, European Union member states, and the United States, compared with the studies reported the occurrence of aflatoxins in date palm fruits.Over the period under review, in studies investigating the levels of aflatoxins contamination in dates in several parts of the world, the finding incidences were compared with the legislation of other international institutions and organizations, such as the European Commission (EC) and the US Food and Drug Administration (FDA). For example, studies conducted in Pakistan indicated that there is no regulation for maximum acceptable levels of aflatoxins in dry fruits; thus, the incidence of aflatoxins contaminated dates was compared with the recommended EC limits for AFB1 and total Afs, respectively [38].


## 5. Discussion

Dates are among the oldest cultivated fruits in the world and are mentioned in the holy texts of Judaism, Christianity, and Islam. Dates thus have religious and cultural values in addition to their nutritive values [47]. Dates consumption is not limited to date production areas [21], and the last ten years have seen a dramatic increase in date imports globally (Figure 1) [23]. Thus, dates are one of the most traded commodities worldwide. Recently, the Food and Agriculture Organization (FAO) approved a request from Saudi Arabia to designate 2027 as the International Year of Dates. The request was initiated because of the Earth’s changing climate and the potential effects on global food security; dates represent a sustainable agricultural commodity grown under various climatic conditions [47].Consequently, monitoring the safety of date fruits regarding aflatoxins contamination is considerably important due to the health risks associated with human exposure to aflatoxins. In this context, strict adherence to guidelines to minimise toxigenic fungi growth and further aflatoxins contamination should be implemented. These guidelines include good agricultural practices during production, good hygienic practices during postharvest handling, and good manufacturing practices during processing [48]. Recently, the use of nanotechnology innovations has been reported to be an effective technique for preserving date palms. For example, the use of edible films and coatings made from chitosan nanoparticles as antifungal materials has been proven to improve the storability of fruits, including date palms [49, 50].The review shows the different analytical procedures (LC, HPLC, LC-MS/MS, and ELISA) utilised to determine the levels of the four targeted analytes (AFB1, AFG1, AFB2, and AFG2) in date palm fruits (Table 1). To detect and quantify aflatoxins without matrix interference from the investigated foodstuff, the toxins should be extracted from the samples and cleaned up, whether chromatography or immunoassay techniques were applied [51]. In the current review, the least common methods of analysis for aflatoxins used were LC and ELISA. Chromatographic methods are among the older analytical techniques used for this purpose. These methods include the LC technique, which is highly sensitive but slower than other methods [52]. On the other hand, ELISA for the analysis of aflatoxins is considered an alternative immunochemical approach to chromatography analysis due to its rapidity, simplicity, and sensitivity. These features are attributed to antibody–antigen interactions, but one of the possible drawbacks of the ELISA technique is its cross-reactivity towards other related toxins [53]. Currently, the combination of LC with fluorescence detection (LC-FD) or with mass spectrometry (LC–MS) or with tandem mass spectrometry (LC–MS/MS) are the most used methods for the quantification of aflatoxins [7]. In the case of the LC-FD method, samples are commonly extracted with methanol or mixtures of methanol and water. Then, an immunoaffinity column (IAC) can be used for clean-up before separation with LC, post-column derivatization, and quantification by FD. Unlike LC-FD, LC–MS and LC–MS/MS do not require derivatisation. For analysis using LC–MS or LC–MS/MS techniques, the samples are commonly extracted with acetonitrile and sometimes in mixtures with water or formic acid before analysis. Nowadays, LC-MS/MS have gained a great popularity and have been widely applied in aflatoxins analysis due to their suitability for multiple toxins detection in a single run with high reliability and sensitivity [7, 54, 55].As shown in Table1, the reported LODs varied according to the diversity of the techniques utilised. However, the indicated LODs of the methods were significantly below the legal limit set by the FDA in the USA, the EU in the European Union, and the GSO in the Gulf States (Table 2). Therefore, it can identify aflatoxins in the investigated date palm fruits that do not comply with current aflatoxins legislation. The susceptibility of dates to aflatoxins contamination over the period under review has been documented at various levels in different world regions. In general, the occurrence of aflatoxins in date fruits was frequently investigated in Pakistan. This could be explained by the fact that Pakistan is one of the major producers of date fruits. On the other hand, no or limited information exist regarding the levels and incidence of aflatoxins in dates from other producers countries, such as Egypt, Saudi Arabia, and Iraq, where dates are highly produced and consumed [19, 22]. The highest documented level of aflatoxins contamination in dates was 26.60 μg/kg among 42.9% positive samples collected from southern areas of Punjab and Khyber Pakhtunkhwa, Pakistan [21]. This level is above the legal limits in the EU for AFB1 and total AFs (2 and 4 μg/kg, resp.) and exceeds the maximum acceptable level in FDA regulation for total AFs (20 μg/kg) (Table 2). In Europe, no incidence of aflatoxins contamination in dates has been reported. This finding could be due to the restrictive regulations for permissible limits for aflatoxins in food, in addition to the availability of effective safety systems, for example, the Rapid Alert System for Food and Feed (RASFF) of the European Union, which seeks to protect human health from food and feed-related risks [56]. Williams et al. [10] commented that the least polluted foodstuff is exported to the developed countries, and the most polluted foodstuff is retained back home to be consumed by the local population in developing countries. Thus, the potential risk of human exposure to aflatoxin varies on a global scale.Heshmati et al. [37] investigated the potential risk of human exposure to aflatoxins in Iran. The results demonstrated that the estimated average daily dietary exposure to AFB1 from dates consumption was 0.12 ng/kg bw/day compared with 0.04, 0.04, and 0.06 ng/kg bw/day for mulberries, figs, and apricots, respectively. In other words, the estimated AFB1 intake from dates consumption was higher than that for other dried fruit. This indicates the fact that the dates can be a likely contributor to the dietary intake of aflatoxins. However, the data available on the contamination of dates by aflatoxins is limited because dates are less frequently investigated compared with other types of foodstuffs.As stated earlier, the recent risk assessment of aflatoxins by the EFSA reported that, among all types of aflatoxins, AFB1 was the most frequent contaminant detected in food, and the importance of continuously monitoring the occurrence of aflatoxins is emphasised due to the Earth’s changing climate and its impact on the increased risk of aflatoxins contamination [7]. Consequently, the risk of human exposure to this carcinogenic component increases and it is classified as a Group 1 carcinogen based on strong evidence of its genotoxicity, which results in the formation of DNA adducts causing point mutations in the TP53 gene [12]. This situation raises concerns regarding the risk of human exposure to aflatoxins through the consumption of dates especially in date-producing regions, such as the Middle East, North Africa and the Arabian Peninsula, where dates are considered as a staple food by the majority [19, 22].


## 6. Conclusion

In conclusion, the current review has presented and discussed the available data regarding the levels and incidence of aflatoxins in dates worldwide, for the first time. This information has clear implications regarding the importance of monitoring aflatoxins contamination in dates. Moreover, the review has highlighted the lack of related data, as the situation is unclear for most regions. Thus, it is important to conduct more comprehensive studies for the occurrence and contamination levels of aflatoxins in dates.



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*Source: 1326861-2022-03-07.xml*