Assessment of Zinc, Iron, and Microbes concentrations in Blood Cockles (Anadara granosa) as Complementary Foods and Implications for reducing of Micronutrition Deficiency

 

Margaretha Solang*, Djuna Lamondo, Ramli Utina, Syam Kumaji, Zuliyanto Zakaria

Department of Biology, Faculty of Mathematics and Natural Science, Universitas Negeri Gorontalo, Indonesia.

*Corresponding Author E-mail: m.solang@hotmail.com

 

ABSTRACT:

Blood cockles (Anadara granosa) are filter feeder that can accumulate heavy metals and microbes so that need to pay attention to the maximum intake. The objective of this study was to evaluate the content of Fe, Zn, a total of microbes and maximum tolerable intake value (MTI). The study employed quantitative analysis. The samples were collected from Wonggarasi Timur Village, Pohuwato Regency, Gorontalo, Indonesia. The determination of Fe and Zn was carried out through Atomic Absorption Spectrophotometer and the microbes total was done by using the Total Plate Count test. Maximum Tolerable Intake value was calculated according to the provisions of JEFCA. The results of the study showed that the Fe and Zn average of blood cockles ranged from 227.85±5.04 ppm – 247.98±6.77ppm and 2.77±0.07ppm-2.82±0.09ppm. Microbes total ranged from 1.7 x 103 colonies/g – 2.4 x 103 colonies/g. The MTI value assumed for person weighing 70 kg based on Fe and Zn content of blood cockle is 0.226kg/day - 0.467kg/day and 24.822kg/day - 24.911kg/day. Blood cockles are a source of Fe, Zn and are safe for consumption according to MTI. Consumption of blood cockles helps reduce the problem of deficiency of Fe and Zn. The content of Fe is higher than the content of Zn in blood cockles. Blood cockles from Pohuwato regency are safe for consumption according to MTI. The microbes total in blood cockles ranged from 1.7x103 colonies/g -2, 4x 103 colonies/g. This total is below the Badan Pengawas Obat dan Makanan/ Indonesian National Agency of Drug and Food Control (BPOM) standard, which is 5 x 105 CFU/g so that the blood cockles are safe for consumption.

 

KEYWORDS: Heavy Metals, Anadara Granosa, Microbes, Safety of Consumption, Gorontalo.

 

 


INTRODUCTION:

Blood cockles (Anadara granosa) or locally known as "bia dara" (in Gorontalo language) is a bivalve group consumed by the people of Wonggarasi Timur Village in Pohuwato Regency, Gorontalo Province, Indonesia. Most of the people of Boalemo and Pohuwato Regency make blood cockles as one of the economically valuable fishery products. Blood cockles have red blood pigment (haemoglobin)1, which is called bloody cockles so that these cockles can live in habitats with have relatively low oxygen levels2,3,4; even after harvesting they can still live without water5

 

Blood cockles are one type of bivalve that can accumulate heavy metals. The heavy metal content found in blood cockles is partly a metal that plays an important role like iron (Fe) and zinc (Zn)6,7,8 and is harmful to human metabolism if its concentration is high. The metal content of cockles varies greatly depending on species, gender, age, and habitat. The Iron level that found in the coastal beach area of Central Tapanuli Regency, North Sumatera, Indonesia reaches 15.1mg/kg9. Furthermore, iron and zinc level of molluscs that found in the southeast coast of India reach 25.65 ppm and 7.7ppm respectively10

 

Iron is in haemoglobin (Hb). Haemoglobin in the blood carries oxygen from the lungs to all body tissues and brings back carbon dioxide from all cells to the lungs to be removed from the body. Iron also plays a role in immunity in the form of lymphocyte cells. Zinc is the second most abundant mineral after iron in the human body. Iron and zinc from food ingredients of these animals do not contain phytate so that they are easily absorbed compared to those derived from plant-based foods. Iron and zinc are essential metals that are needed by the body in small amounts. On the other hand, Iron (Fe) and zinc (Zn) are essential elements needed by the body, acting as constituents of enzymes and other essential proteins involved in the metabolic cycle.

 

On the other hand, given that blood cockles are filter feeders so that these cockles can have the ability to accumulate microbes. The microbial aspect has a significant role in the safety of marine products. Study of Fe and Zn content of blood cockle in Boalemo District, Gorontalo Province, Indonesia has been carried out but has not revealed the safe limit of consumption of these blood cockles. Meanwhile, the potential for blood cockles from Pohuwato Regency and the safety of their consumption have not been studied. This study was conducted to evaluate the content of Fe, Zn, microbes total, and Maximum Tolerable Intake Value (MTI) of blood cockles in Pohuwato Regency, Gorontalo, Indonesia. Calculation of Maximum Tolerable Intake required reducing the risk for the health of people who consume blood cockles.

 

MATERIAL AND METHODS:

Study area

The samples of blood cockles were taken from Wonggarasi Timur village of Pohuwato regency, Gorontalo Province, Indonesia. Sampling was carried out between June and July 2017, at low tide at 5.30 pm. The samples were frozen in ice and stored in the icebox. The sampling location consists of three sites in Wonggarasi Timur village. A site 1 is located in the mangrove area, site 2 is in 200m of the mangrove area, and site 3 is in the estuary area (Figure 1). These three locations were chosen because this location is where blood cockles live. The results of temperature measurements at three sites are as follows, the site 1: 27ºC, site 2: 28ºC, and site 3: 27ºC. The results of measuring water salinity at three sites were 32.5 0/00 (a site 1), 32.1 0/00(site 2) and, 31.7 0/00 (site 3).

Sample preparation:

Before being analyzed, blood cockles’ samples were cleaned with running water from the tap to remove sand particles. Blood cockle meat was separated from the shell and washed with running water. All samples were made in triplicate and stored in an oven at a temperature of 60ºC until a constant weight was obtained11.

 

Sample analysis:

Iron and zinc were measured by Atomic-Absorbent Spectrophotometric (AAS) ZEEnit 700 brand. The Fe and Zn analysis were conducted at the Balai Besar Kesehatan Daerah Surabaya (Surabaya Health Institution), Indonesia. The Fe and Zn were measured in ppm. Determination of the number of microbes is based on the method through the Total Plate Count (TPC) test and is carried out in the Biology Laboratory of the Faculty of Mathematics and Sciences (FMIPA-Indonesia.

 

Data analysis:

Data from the results of sample analysis of Fe, Zn, and averages of microbestotal were averaged; while the safety analysis of blood cockle consumption was done by calculating the Maximum Tolerable Intake (MTI) according to the provisions of JEFCA in 1982 and 198312. The method of calculating MTI is as follows:

 

MTI = Estimate PMTDI / Ct

 

Notes:*) MTI: Maximum Tolerable Intake*) Estimate PMTDI: Estimated value of Provisional Maximum Tolerable Daily Intake (mg/day)*) Ct: The concentration of heavy metals found in cockles soft tissue (mg/kg). Also, the Fe and Zn data obtained were compared with the maximum allowable metal content of clams (Decree of the Directorate General of Drug and Food Control No 03725/B/SK/VII/89). Total of microbes data was determined based on the average analysis results and compared to the Indonesian National Standard.


 

 

Figure 1. Blood cockles sampling sites


RESULTS AND DISCUSSION:

The level of iron (Fe) in blood cockles:

The results showed that blood cockle which is a group of molluscs have potential as a source of Fe. Iron (Fe) is an important metal that plays a role in the formation of red blood cells so that molluscs can be recommended for consumption by pregnant women and children13. Fe content found in blood cockles for each location was different. Blood cockles’ Fe content is shown in Table 1. The content of Fe in the blood cockles was found to be very high. The presence of Fe content in blood cockles was assumed to be due to blood cockles as benthic organisms (dwelling organisms) that are filter feeders. The average levels of blood cockles’ Fe found in the site 1, 2, and 3 respectively are 247.98±6.77ppm, 228.79± 6.71ppm, and 227.85±5.04ppm.

 

Table 1. The levels of Fe (ppm) in blood cockles were taken from Wonggarasi Timur village of Pohuwato Regency, Gorontalo Province, Indonesia.

Observation site

repeat

average (ppm)

SD

 

1

2

3

 

 

1.

259.67

244.16

244.85

247.98

6.77

2

225.85

223.70

238.65

228.79

6.71

3

233.44

223.65

226.47

227.85

5.04

 

The highest Fe content was found at the site I, and the lowest Fe content was found at site III. This high level of Fe is assumed to be related to blood cockles habitat. There is human activity around the blood cockles habitat found which contributes as a source of heavy metals14. Another factor which is the source of heavy metals in Pohuwato waters between housing, landfills and markets.15,14 The presence of heavy metals can accumulate in the body of biota around sediments such as blood cockles. Around the mangrove area (a site I) is an area close to garbage disposal, markets, and community mining. While at site III is an area near the settlement.

 

The Iron content of blood cockles from this study was higher than blood cockles’ Fe content found in Boalemo Regency (93.91 ppm)6 and Kuala Selangor, Malaysia (25.22mg/kg)16. However, the Fe blood cockle content of Pohuwato is lower than the Fe content of blood shells found in Kuala Selangor, Malaysia, which is around 875.55±396.55mg/kg14 and in the waters of Tanjung Mas, Semarang, Central Java, Indonesia and the waters of the Wedung, District of Demak, Central Java, Indonesia, which averaged between 563.32-1, 305.05 mg/kg wet weight7. (Table 2).

 

 

Table 2. The level range of Heavy Metals Fe in Blood cockles

Studies

Location

Level range

This recent study

Pohuwato Regency, Gorontalo Province, Indonesia.

227.85 ± 5.04 ppm – 247.98 ± 6.77 ppm

Nurjanah et al., 2005

Boalemo Regency, Gorontalo Province, Indonesia

93.91 ppm

Ishak et al., 2016

Kuala Selangor, Malaysia

25.22 mg/kg

Yunus et al, 2014

Kuala Selangor, Malaysia

875.55 ± 396.55 mg/kg

Taurusiana et al, 2014

in the waters of Tanjung Mas Semarang and the waters of the Wedung, Demak, Province of Central Java, Indonesia

563.32 - 1,305.05 mg/kg

 

Blood cockle is classified into molluscs. Molluscs are a good source of minerals so consuming molluscs will help reduce the problem of micronutrient deficiency. Clam found in Rivers State, Nigeria is reported to contain 25.71mg/g Fe17. High levels of Fe in blood cockles are used as dietary supplements for anemia patients18.

 

The levels of Zinc (Zn) in blood cockles:

Zinc is a heavy metal needed by the body. Zinc plays an important role in cell growth, differentiation, and metabolism. Zn deficiency can interfere with growth and decrease immunity12. Zn supplementation improves the growth of infants and children experiencing growth disorders19,20.

 

The results showed that the average level of Zn in blood cockles found in the site 1, 2, and 3 is 2.82±0.09ppm; 2.77±0.07ppm; and 2.81±0.03ppm (Table 3). Zn levels at three observation sites were different. This was considered because of community activities in these three different observation sites. The first site (Site 1) is a place for ship welding, garbage disposal, and fish auctions. This community activity produces waste that can enter the waters, then settles on the sediments and can accumulate in the body of the biota, especially the clams that live in it.

 

Tabel 3. Levels of Zn (ppm) in blood cockles were taken from Wonggarasi Timur village of Pohuwato regency, Gorontalo Province, Indonesia.

Observation site

repeat

average (ppm)

SD

 

1

2

3

 

 

1.

2.96

2.81

2.70

2.82

0.09

2

2.87

2.72

2.72

2.77

0.07

3

2.838

2.78

2.81

2.81

0.03

 

 

The Zn levels of blood cockles in this study were below the standard limit of Zn metal in marine biota muscles, namely 100mg/kg21,22. Zn levels in blood cockles found in this study were lower than those of blood cockles’ Zn found in blood cockles (13.91ppm) from Boalemo Regency, Gorontalo, Indonesia and Kuala Selangor, Malaysia (3.12mg/kg)16, and Kuala Selangor, Malaysia (94.17±9.13mg/kg)14, and in the waters of Tanjung Mas, Semarang and the waters of the Wedung, Demak, Indonesia averaged range 563.32-1, 305.05mg/kg (Table 4). This study shows that blood cockles from Pohuwato tend to accumulate Fe higher than Zn. The ability of blood cockle to accumulate Fe higher than Zn is also seen in blood cockle found in Kuala Selangor, Malaysia14,16. The results of this study also show that clams which are a group of molluscs are a source of metals Zn needed for the body17. Blood cockle supplementation increases serum Zn levels in the Rattus norvecigus malnutrition22.

 

Table 4. The level range of Heavy Metals Zn in Blood cockles

Studies

Location

Level range

This study

Pohuwato Regency, Gorontalo Province, Indonesia.

2.77 ± 0.07 ppm - 2.82 ± 0.09 ppm

Nurjanah et al., 2005

Boalemo Regency, Gorontalo Province, Indonesia

13.91 ppm

Ishak et al., 2016

Kuala Selangor, Malaysia

3.12 mg/kg

Yunus et al., 2014

Kuala Selangor, Malaysia

94.17 ± 9.13 mg/kg

Taurusiana et al., 2014

in the waters of Tanjung Mas Semarang and the waters of the Wedung, Demak, Province of Central Java, Indonesia

68,13 – 94,22 mg/kg

 

Total of Microbes:

Seafood products are usually consumed immediately after being caught. However, contamination by pathogenic bacteria can occur during capture, handling and processing. Aquatic products such as fish, shrimp, clams, and others have great potential as a cause of food poisoning. The results of the analysis of the total of blood cockle microbial taken 200 m from the mangrove and at the estuary respectively were 1.7x103 colonies / g; 2.1 x 103 colonies / g, and 2, 4 x 103 colonies/g. The highest total of microbes was found in the observation point I area around mangrove. The lowest total of microbes is found in cockles taken from areas within 200 m of mangrove. If it is seen from the total of microbial values in fresh blood cockles, it can be said that the samples are still microbiologically safe.

 

Blood cockles have a moisture content of around 74.37% 6. This moisture content is high, so it is susceptible to damage by microbiologists23. The number of blood cockles microbes from Pohuwato Regency is under the SNI standard No. 7388: 2009; BPOM, 2009, which is 5 x 105 colonies/g24,25. The number of microbes found in blood cockles shows that blood cockles products are safe from biological microorganisms. It also shows that sanitation conditions during processing are good. However, the presence of microbes in blood cockles also shows the need for better handling during transportation from fishermen to consumers, for example, by cooling or good sanitation. Improper handling can cause microbes to grow and multiply so that blood cockles are no longer safe to consume.

 

The consumption safety based on Fe and Zn levels of blood cockles:

Foods containing heavy metals can be harmful to the body when consumed. The maximum tolerance limit for consumption of food containing heavy metals has been determined using consumption safety figures. The Joint FAO/WHO Expert Committee on Food Additives (JECFA) in 1982 and 1983 established consumption safety figures called Maximum Tolerable Intake (MTI) or more commonly called maximum tolerance consumption. The value of MTI for blood cockles based on Fe and Zn levels found in this study is assumed for 70 kg body weight that can be seen in Table 5.

 

The maximum tolerance for metal (Fe) has been determined by JEFCA (1983) of 56 mg per day for people weighing 70 kg. If the amount of iron that enters the body of a person weighing 70 kg exceeds this value, then the metal Fe will be toxic in the body. Fe content in blood cockles found in mangrove area was 247.98 ppm. A person weighing 70 kg can consume blood cockles meat no more than 0.226 kg/day or 226 g/day. Whereas for blood cockles are taken from areas with a distance of 200 m from mangroves can be consumed no more than 0.245 kg/day or 245 g/day. While for blood cockles are taken from areas near the river mouth can be consumed no more than 0. 467 kg/day or 467 g/day.

 

The daily requirement of Zn for adults is around 15 mg per day. The maximum tolerance consumption for Zn metals has been determined by JEFCA (1982) of 21-70 mg per day for people weighing 70 kg. If the amount of Zn entering the body with a bodyweight of 70 kg exceeds this value, the Zn metal will be toxic in the body. Zn levels in blood cockles found on Wonggarasi beach mangrove area ranged from 2.77-2.82 ppm so that people weighing 70 kg can consume blood cockles between 7.447–24.822 kg/day. Whereas for blood cockles are taken from areas with a distance of 200 m from mangrove area can be consumed no more than between 7.581–25.277 kg per day. While for blood cockles taken from areas near the river mouth can be consumed no more than 7.473 – 24.911 kg/day.

 

The average number of blood cockle consumption in the Pohuwato community, Gorontalo, Indonesia in this study is based on JEFCA, this is different from the average blood cockle consumption from Kuala Selangor, Malaysia. The value of blood cockle consumption from Kuala Selangor is based on the average value of blood cockle consumption in the Malay, Chinese and Indian ethnic communities in Peninsular Malaysia [14]. Based on the content of Fe blood cockle, the estimated daily intake of Malay, Chinese and Indian people is 4692.3 g/day, 2714.19 g/day, and 3160.72 g/day. Meanwhile, if based on Zn content of blood cockle, the people of Malay, China and India can consume blood cockle as follows, 339.97 g/ day, 291, 94 g/day, and 339.97 g/day.

 

Table 5. Maximum Tolerable Intake of blood cockles (JECFA)

Metals

PMTDI (mg/kg/bd/day) (a)

Estimate PMTDI (mg/day) (b)

MTI (c) (kg/day)

Observation site (This study)

I

II

III

Fe d

0.8

56

0.226

0.245

0.461

Zn e

0.3 -1

21-70

7.447 - 24.822

7.581 - 25.277

7.473 - 24.911

Note: a). PTMDI = Provisional Tolerable Maximum Daily Intake b). For people weighing 70 kg c).

MTI = Maximum Tolerable Intake d). JEFCA (1983) (WHO,2019) e). JEFCA (1982) (WHO, 2019)

 

CONCLUSION:

The Iron (Fe) content of Wonggarasi beach blood cockles in Pohuwato Regency, Gorontalo province ranged from 227.85 - 247.98 ppm, while blood cockles’ Zinc levels ranged from 2.77 ppm - 2.82 ppm. The results of this study indicate that the content of Fe is higher than the content of Zn in blood cockles. Maximum Tolerable Intake (MTI) assuming that for 70 kg body weight according to JEFCA (1983) for 247.98 ppm Fe is 0.226 kg or 226 grams per day, whereas for blood cockles with 228.79 ppm Fe can be consumed no more than 0.245 kg/day or 245g/day. Meanwhile for blood cockles with Fe 227.85 ppm taken from areas near the river mouth can be consumed no more than 0. 467 kg/day or 467 g/day. Maximum Tolerable Intake (MTI) with the assumption for people with a bodyweight of 70 kg according to the provisions of JEFCA for Zn metals with a level of 2.82 ppm is 7,447–24,822 kg/day. Whereas for blood cockles containing 2.77 ppm Zn can be consumed no more than between 7,581-25,277 kg/day. While for blood cockles containing 2.81 ppm Zn can be consumed no more than 7,473-24,911 kg/day. Blood cockles from Pohuwato regency are safe for consumption according to MTI. The microbes total in blood cockles ranged from 1.7 x 103 colonies/g - 2, 4 x 103 colonies/g. This total is below the BPOM standard, which is 5 x 105 CFU/g so that the blood cockles are safe for consumption.

 

ACKNOWLEDGEMENT:

The author would like to thank the Institute of Research and Community Service of Universitas Negeri Gorontalo, Indonesia for funding this research.

 

CONFLICT OF INTEREST:

The authors declare no conflict of interest.

 

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Received on 01.06.2020           Modified on 02.07.2020

Accepted on 26.07.2020         © RJPT All right reserved

Research J. Pharm. and Tech. 2021; 14(6):3399-3403.

DOI: 10.52711/0974-360X.2021.00591