2. IARC report on the carcinogenicity of chewable tobacco & Betel quid (Tambula / Pan) in 1985

 

The International Agency for Research on Cancer (IARC), an intergovernmental agency affiliated to World Health Organization (WHO) of the United Nations (UN) published its first report on areca nut in 1985 in the for of a monograph (Vol. 37). This monograph was an outcome of a one-year effort involving 19 experts drawn from 10 countries (working group), 5 observers from 5 countries and 22 secretaries affiliated to IARC. After reviewing and analyzing numerous research papers published in reputed research journals, a week-long workshop at Lyon, France, the experts came out with a comprehensive report in the form of a monograph in 1985.

The working group looked into the carcinogenicity chewable tobacco, betel quid & areca nut in the consumers. The experts looked into the reports published on the evidence of carcinogenicity in experimental animals, DNA damage studies, Cell-line studies, case reports involving human subjects & epidemiological studies.

Based on these studies and a systematic evaluation, IARC came out with the following conclusions with respect to chewable tobacco, betel quid and areca nut as follows.

Chewable tobacco

1. There is sufficient evidence that oral use of chewable tobacco is carcinogenic to humans. 
2. There is sufficient evidence that oral use of tobacco mixed with lime (khaini) is carcinogenic to humans.

Betel quid (Pan / Tambula) comprising areca nut, betel leaf, slacked lime, catechu.

1. There is sufficient evidence that the habit of chewing betel quid containing tobacco is carcinogenic to humans.
2. There is inadequate evidence that the habit of chewing betel quid without tobacco is carcinogenic to humans.
3. The Working Group also concluded that, while there is sufficient evidence that the combined habits of smoking tobacco and chewing betel quid without tobacco cause oral and pharyngeal(throat) cancer.
4. There is limited evidence that areca nut with and without tobacco is carcinogenic to experimental animals.
5. The data are inadequate to allow an evaluation of the carcinogenicity of betel leaf or arecoline to experimental animals.

As per this monograph, neither areca nut nor the other constituents of betel quid / Tambula/ Pan (areca nut, betel leaf, Catechu and slacked lime) were implicated for carcinogenicity.  

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1. Prominent reports on carcinogenicity of Areca nut & Betel quid (Pan / Tambula)

 

More than 50 research papers were published during 1970 to 1980 on the link between Betel quid / Areca nut along with tobacco chewing and mouth & throat cancer. Many reports pointed out that the percentage of oral cancer among all cancers diagnosed in hospitals in Asia was always much higher than that usually found in western countries. In western countries, the habit of chewing betel quid, with or without tobacco, is virtually unknown. In many descriptive studies, investigators have obtained histories of chewing betel quid with tobacco from series of patients with oral cancer. In most of these studies, the percentage of patients who practice chewing habits is extremely large. Many research papers have revealed that a high incidence of oral, oro- and hypopharyngeal cancer is observed in regions of the world where a high proportion of the population practices betel-quid chewing. Out of the 2,67,000 new oral cancers estimated to occur in the year 2000 globally, 1,28,000 (48%) occur in South and South-East Asia. Out of 1,23,000 cases of oro- and hypopharyngeal (throat) cancer estimated to occur globally annually, 63 000 (51.2%) are accounted in South and South-Easts Asia as per an assessment by Ferley & coworkers of International Agency for Research on Cancer (IARC).

The International Agency for Research on Cancer (IARC) is an intergovernmental agency affiliated to World Health Organization (WHO) of the United Nations (UN). IARC has its own governing council, comprising 27 member countries including India as on today. Almost all the developed nations of the world are the members of the governing council of IARC.

IARC was stablished in 1965 by the World Health Assembly, as an autonomous agency of WHO with the aim of promoting international collaboration in cancer research. Its mission is to coordinate international studies on the causes of human cancer, the mechanisms of carcinogenesis and strategies for cancer prevention, with a particular focus on promoting research in regions of the world where it is lacking. As IARC is affiliated to WHO independent of any government, there is a belief that it provides a reliable and authoritative assessments of many facets of cancer information valued by scientists, governments, nongovernmental organizations and the general public, globally.

The evaluations of carcinogenic risk are made by international working groups of independent scientists. A detailed account of the procedures involved, research papers referred and methodology followed for analysing the data etc. are given in the monograph. Sufficient evidences are provided by IARC that complete transparency in the process of monograph preparation is followed. The monographs are prepared based on the examination of all relevant information (research papers published in reputed journals) in order to assess the strength of the available evidence that certain exposures could alter the incidence of cancer in humans. Based on thorough study, the agents/substances are classified as follows.

       i.         Group 1: The agent is carcinogenic to humans. Ex.: Lindane, soot, tobacco smoke, Benzene.

     ii.         Group 2A: The agent is probably carcinogenic to humans. Ex.: Malathion, Glyphosate (a popular weedicide).

   iii.         Group 2B: The agent is possibly carcinogenic to humans. Ex.: Chloroform Chlorophenoxy herbicides, DDT, Aloe vera whole leaf extract.

    iv.         Group 3: The agent is not classifiable as to its carcinogenicity to humans. Ex.: Ampicillin, Caffeine, Eugenol, Tea.

     v.          Group 4: The agent is probably not carcinogenic to humans.

So far, IARC has published four reports on the carcinogenicity of areca nut and its components, chewing of tobacco with or without betel quid (Tambula/Pan), with or without areca nut, as follows.

1.     Evaluation of the Carcinogenic risk of tobacco, Betel quid and areca nut chewing. Published in 1985, Vol. 37.

2.   Evaluation of carcinogenic risks to humans from betel-quid and areca-nut chewing and some areca-nut-derived nitrosamines. Published in 2004, Vol. 85.

3.  Evaluation of Carcinogenic Risks to Humans from Smokeless Tobacco and Some Tobacco-specific N-Nitrosamines. Published in 2007, Vol. 89.

4.      Identification of carcinogenic hazards from arecoline. Published in 2021, Vol.128

A detailed commentary along with critical comments shall be provided on each report in my future Blogs.

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9. Alkaloids present in Areca nut - Part IV

 

In the last couple of Blogs, the arecoline & other alkaloid contents in the green immature areca nut studied in different countries was discussed. In this Blog, arecoline & other alkaloid content of the areca fruits of 9-10 month’s maturity is discussed.

Shivashankar et al., (1968) from CFTRI Mysore, had studied the chemical composition of Chali (White supari) purchased from important areca growing regions of Kerala and Karnataka over two years (1964 to 1966). They had used rudimentary methods available at that time to estimate arecoline content. They concluded that the arecoline content varies between 0.1 to 0.7 % in Chali on dry weight basis, collected from different areca growing regions of Karnataka and Kerala state of India. 

Wang et al., (1997), had studied the arecoline content of areca nuts of 9-10 months’ maturity collected from Taiwan. They had reported the arecoline content of 0.16% in the ripened fruit on fresh weight basis. Jantarat et al., (2013) had reported an arecoline content of 0.094 % in 8 months old, matured nuts on fresh weight basis. They had collected the areca nut sample from Nakhon Si Thammarat, Thailand.

Chavan and Singhal (2013) had analyzed the arecoline content of Chali purchased from the local market in Mumbai using HPLC method. They had reported an arecoline content of 1.279% (12.79 mg/g).  Jain et al., (2017), have done a comprehensive study of arecoline content in Chali and various areca nut products sold in the market in China and India. They had reported an arecoline content of 0.064 to 0.222 % (dry weight) using advance LC-MS/MS method. Arecoline content of 0.119% was the median value and 0.127% was the mean value in the Chali (white supari) collected from the bulk traded in the Mumbai market. The arecoline content found in the Gutka samples produced and sold in India were 0.074 & 0.116 %. The arecoline content in the Bing lang purchased from Hunan province China were in the range of 0.107 to 0.144%.  Levels of Guvacine ranged from 0.139 to 0.816% dry weight, accounting for more than 50% of the total alkaloid content in most of the products. The levels of Arecaidine and Guvacoline ranged from 0.014 % to 0.170 % and 0.017 % to 0.099 % dry weight, respectively.

In conclusion, about a dozen alkaloids have been identified so far in areca nuts and Arecoline is considered as a key biologically active alkaloid among them. Its concentration will be high in the early developmental stage and decreases with maturity. Areca nut cultivated in Taiwan, China, Indonesia and Darwin Island, Australia, reported arecoline content on higher side (0.3% and above) in the unripe green nuts. Whereas the unripe green areca nuts studied in Thailand and India had lower arecoline content (less than 0.3%). Lot of variation (up to 400%) in arecoline concentration was found in areca nuts cultivated in different regions.  The analysis of various areca nut samples sold in the market revealed that the arecoline content in them is in the range of 0.064 to 0.222 % (0.64 to 2.22 mg/g). In one instance, the arecoline content was found to be as high as 1.279% (12.79 mg/g) in Chali sample collected from Mumbai market, India.

References:

Shivashankar et al., (1968). Physical and chemical characteristics of processed areca nuts. Journal of Food Science & Technology, Vol.  6, pp. 113-116.

Wang et al., (1997), Contents of Phenolics and Alkaloids in Areca catechu Linn. During Maturation, Journal of Agriculture & Food Chemistry. Vol. 45, pp. 1185-1188

Jantarat et al., (2013), A Simple and Rapid HPLC Technique for Determination of Arecoline in Areca Nut (Areca catechu L.) Extract, Walailak Journal of Science & Technology, Vol. 10(1), pp. 57-66.

Chavan & Singhal (2013). Separation of polyphenols and arecoline from areca nut (Areca catechu L.) by solvent extraction, its antioxidant activity, and identification of polyphenols. Journal of  Science of  Food & Agriculture.  Vol. 93(10), pp. 2580–2589.

Jain et al (2017). Analysis of Alkaloids in Areca Nut-Containing Products by Liquid Chromatography−Tandem Mass Spectrometry. Journal of Agricultural and Food Chemistry, Vol. 65, pp. 1977−1983

8. Alkaloids present in Areca nut - Part III

In the last two Blogs, the arecoline content of green areca nuts of 6-7 months maturity has been discussed. In the current Blog, Arecoline and other alkaloids content of green nuts of 2-5 months maturity is being discussed. Areca nuts of this maturity is being utilized for consumption in Taiwan and China.
Wang et al., (1997), had studied the arecoline content of fresh areca nuts of 2 and 3 months’ maturity collected from Taiwan. They had reported the concentration of all the alkaloids in 2-3 months’ maturity as follows.

 

Yuan et al., (2012) had studied the arecoline content of fresh areca nuts collected from twelve prominent areca growing districts of Hainan province of China and had analyzed those fresh nuts (perhaps 4-5 months’ maturity). They had also estimated the concentration of Arecaidine and Guvacine in them as follows.

Jantarat et al., (2013) had reported an arecoline content of 0.143 % in 3 months old fresh nuts.  They had purchased the areca nuts from Nakhon Si Thammarat, Thailand and had used HPLC to estimate the alkaloids. 

From the above cited reports, it is clear that the arecoline content in the green nuts of Taiwan (2-3 month's maturity) was in the range of 0.7 % (7 mg/g), whereas the arecoline content was mere 0.143% (1.43 mg/g) in the areca nut from Thailand. A significant variation was found in the arecoline content in the green nuts collected from Hainan China as well. The values were ranging between 0.217 to 0.764 % (2.17 to 7.64 mg/g)

References:

1.    Wang et al., (1997), Contents of Phenolics and Alkaloids in Areca catechu Linn. during Maturation, Journal of Agriculture & Food Chemistry. Vol. 45, pp. 1185-1188

2.     Yuan et al. (2012), Alkaloids from Areca (Betel) Nuts and Their Effects on Human Sperm Motility In Vitro. Journal of Food Science , Vol. 77 (4), pp. T70-78.

3.     Jantarat et al., (2013), A Simple and Rapid HPLC Technique for Determination of Arecoline in Areca Nut (Areca catechu L.) Extract, Walailak Journal of Science & Technology, Vol. 10(1), pp. 57-66.

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7. Alkaloids present in Areca nut - Part II

 

A study was undertaken by our team in NITK Surathkal in 2021 and 2022 to study the arecoline content in the unripe areca nuts of 6-7 months’ maturity. The areca nut samples were collected from various locations as explained in my Blog 4. Phenolic compounds present in Areca nut (Karnataka, India): Part – II (under the label Chemical constituents of Areca nut). The arecoline content of the fresh areca nut samples was determined immediately after the collection. Very sensitive HPLC method was employed to find the arecoline content in the samples. The results are presented below.

The median arecoline content among the 21 samples was 2.17 mg/g on Fresh weight (FW) basis. Sample from K. Hosur, Tarikere Taluk, Chickmagalur District had the highest arecoline content of 3.66 mg/g (0.366 %) on fresh weight basis. Whereas the sample from Gajanur, Shivamogga district had the lowest arecoline content of  0.71 mg/g (0.071%) on fresh weight basis.

In general, the arecoline content of all the samples analyzed in this study was on the lower side compared to the reports published elsewhere. Only three samples had the arecoline content more than 0.3 % out of 21 samples analyzed. In general, if areca nut is having 0.3% arecoline content (3.0 mg/g) is dried as it is, the dried nut would contain 0.75 % (7.5 mg/g) arecoline on dry weight basis. Interestingly, two samples had arecoline content less than 0.1%, which is a remarkable discovery.                                                                                                                                                                                                      

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6. Alkaloids present in Areca nut - Part I

 

Arecoline, arecaidine, guvacine, and guvacoline are the most important and prominent alkaloids found in areca nut. Nicotine, dichroine, acatechu A, acatechu B, homoarecoline, N-ethyl-1,2,5,6-tetrahydro-1-methyl-3-pyridine carboxamide and arecatemines are some of the minor alkaloids found in some areca nuts. Arecoline is found to be the most important alkaloid among all, due to its role in modulating many physiological functions upon human consumption.

In humans, depending on the dose and individual responsivity, arecoline produces cognition-enhancing, psychostimulant, euphoric, pro-arousal, aphrodisiac, anxiolysis and sedative effects. In other words, arecoline produces enhanced alertness and learnability, enhances the mood, gives a feeling of great pleasure, causes awakening, stimulates the sexual desire, reduces anxiety and promote calmness. Arecoline has a wide spectrum of pharmacological effects beyond Central Nervous System modulating cardiovascular, digestive and endocrine systems as well.  Like nicotine, arecoline also evokes addiction and withdrawal symptoms (upon discontinuation). Withdrawal symptoms could be mood swings, anxiety, irritability and insomnia.

Arecoline content in Green Areca nuts (6-7 month's maturity)

Many research groups have studied the arecoline content in areca nuts cultivated in various geographical locations and at various level of maturity. Mathew et al., (1964) from CFTRI Mysore, had studied the arecoline content of areca nuts of 6-7 months maturity collected from Kyatsandra, Mysore, Thirthahalli and Vittal in Karnataka state and Pudupady in Kerala state of India.  They have concluded that the arecoline content was in the range of 0.14 to 0.19 %.

Huang and Mcleish (1989) had reported an arecoline content of 0.3-0.63 % in fresh areca nuts (perhaps 6-7 months’ maturity) collected from Darwin, Australia using HPLC method. This was the first report of use of HPLC for estimating arecoline. They had estimated the arecaidine (0.31-0.66%), guvacoline (0.03-0.06%) and guvacine (0.19 0.72%) in those samples as well.

Sari et al., (2020) had studied the arecoline content of unripe green areca nuts (6-7 months’ maturity) collected from two different regions of Indonesia. They had reported an arecoline content of 0.44 & 0.69 % (on dry weight basis).

References:

1.  Mathew et al. (1964) Studies on Arecanut: Part I - Changes in Chemical Composition & Physical Characteristics of Nuts with MaturityIndian Journal of Technology, Vol. 2, No.3, pp. 90-96

2.    Huang and McLeish (1989). High-performance liquid chromatographic determination of the alkaloids in betel nut, Journal of Chromatography, 415, pp. 447-450.

3.   Sari et al., (2020). Distinct phenolic, alkaloid and antioxidant profile in betel quids from four regions of Indonesia, Scientific Reports, Vol. 10:16254

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