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Research Article
 

Microbial and Heavy Metals Contamination of Herbal Medicines



S.S. Alwakeel
 
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ABSTRACT

This study was conducted to evaluate the microbial contaminants and presence of toxic heavy metals on some herbal medicines. Twenty-seven samples (3 kg each) of well-known herbs and 5 kinds of henna available in herb markets around Riyadh, Saudi Arabia were collected for microbial and toxic metal contamination. Twenty-one (60%) of the samples showed the presence of fungi. Aspergillus flavus and Aspergillus fumigatus were the most common isolates (9/35, 35%). Astralagus sarcocolla had the highest TPC count; Matricaria chamomilia had the highest total coliform count and fecal coliform count. Two henna samples showed more than 1 ppm lead content. Mercury was the highest in Lepidium sativum, aluminum in Zingiber officinale, calcium in Artemisia herba alba, cadmium in Lepidium sativum, Vigna radiata and Zingiber officinale. Copper was the highest in Cinnamomum zeylanicum, iron in Zingiber officinale, zinc in Salvia officinalis, potassium in Matricaria chamomilia, whereas sodium was the highest in Pimpinella anisum. Microbial determination showed that Bacillus species was seen in 3 (9.7%) of the isolated microorganisms with the predominance of Bacillus cereus (14/31, 45.2%). Other microbial isolates were Aeromonas hydrophilia, Shigella spp., Enterobacter agglomerans, Enterobacter spp., Vibrio fluvialis, Escherichia coli, Pasteurella multocida, Enterobacter cloacae, Staphylococcus hyicus, Staphylococcus epidermidis, Acinetobacter iwoffii and Klebsiella. Tests showed the sensitivity of most isolated bacteria to amoxicillin, gentamicin, imipinem, tobramycin and trimethoprim-sulfamethoxazole.

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  How to cite this article:

S.S. Alwakeel , 2008. Microbial and Heavy Metals Contamination of Herbal Medicines. Research Journal of Microbiology, 3: 683-691.

URL: https://scialert.net/abstract/?doi=jm.2008.683.691
 

INTRODUCTION

Herbal drugs have increasingly been used worldwide during the last few decades as evidenced by rapidly growing global and national markets of herbal drugs. According to WHO estimates, the present demand for medicinal plants is about US $14 billion a year and by the year 2050 it would be about US $5 trillion. Now people rely more on herbal drugs because of the high prices and harmful side effects of synthetic drugs and this trend is growing, not only in developing countries but in developed countries too. Unfortunately, the number of reports of people experiencing negative effects, caused by the use of herbal drugs, has also been increasing. There may be various reasons for such problems and one of the major causes of adverse effects is directly linked to the poor quality of herbal medicines. Therefore, it is realised that insufficient attention was being given to the quality assurance and control of herbal medicines. Although WHO has developed guidelines to maintain the quality of herbal drugs which includes a detailed description of the techniques and measures required for the appropriate cultivation and collection of medicinal plants. Despite such guidelines, there is still a lacuna between this available knowledge and implementation, because farmers and other relevant persons like producers, handlers and processors of herbal drugs are not very aware of WHO`s guidelines.

They continue their work, as before without any quality control measures which results in inferior quality of herbal drugs with lots of contaminants like heavy metals, pesticides and microbes.

Several studies showed that herbal plants are associated with a broad variety of microbial contaminants (Kneifel et al., 2002; Ichinoe et al., 1988). Recently, Ang (2003) showed that 22% of herbal plant samples studied failed to comply with quality requirement for traditional medicines. On the other hand, many studies have investigated the presence of toxic contaminants in herbal plants. Trace elements such as zinc, manganese, chromium, copper, iron, lead, nickel and vanadium were found in a traditional Chinese herb Jinqi (Han et al., 2008). In 2007, a 93 year old hypertensive woman was reported with severe hypokalemia due to consumption of licorice-containing herbal medicine for 7 years (Yasue, 2007). Another study showed that 26% of the available herbal plants in Malaysia possessed 0.53-2.35 ppm of mercury and therefore, do not comply with the quality requirement for traditional medicine (Ang and Lee, 2006). A study in Brazil (Caldas and Machado, 2004) showed that samples of herbal medicines had cadmium up to 0.74 mcg g-1 and mercury up to 0.087 mg g-1 and lead estimated intake through consumption of these herbs reached 440% of the tolerable intake. Metal toxicity has been associated with pathophysiological effects which included neurological behavioral effects (Boyd et al., 1991; Echeverria et al., 1998), cardiac dysfunction (Frustaci et al., 1999), fetal malformations (Vimy et al., 1990), Alzheimer`s disease (Cornett et al., 1998) and Parkinson`s disease (Ngim, 1989). The concentration of heavy metals is increasing in the environment and many hazardous effects are caused in the inhabitants of that environment. Modern detection methods have revealed trace amounts of lead in plants and water. However, these herbs have never been shown, or even suspected, of causing any disease associated with lead poisoning. For such reasons, we opted to investigate the microbial and toxic metal contamination of the most common herbal products available from random markets in Saudi Arabia`s capital city of Riyadh.

MATERIALS AND METHODS

Sampling
Twenty-seven samples of well-known plant herbs and 5 different types of henna were collected at random in February 2008 from random herbal markets in the city of Riyadh, Saudi Arabia to determine the predominant microorganism and mycoflora and to assess the toxic heavy metal content. The herbal products were chosen on the basis of their commercial availability and popularity of use (Table 1). Every sample weighed 3 kg and was stored inside clean plastic bags at a temperature ranging from 4-5°C.

Evaluation of Fungal Contamination
Every sample was examined for Total Fungal Count (TFC) according to Nordic Committee on Food Analysis (2005), Total Coliform Count (TCC) according to Nordic Committee on Food Analysis (2004), Fecal Coliform Count (FCC) according to Nordic Committee on Food Analysis (2005), staphylococcus count according to Nordic Committee on Food Analysis (1999) and Bacillus cereus count according to Nordic Committee on Food Analysis (2003). Five grams of each sample were mixed with 45 mL of diluent from which tenfold serial dilution was made. Three milliliter from each dilution was inoculated each in sterile petri dishes in which sterile media was poured. After solidification plates were incubated. Plates were incubated upside down at 26±1°C for 7 days. After incubation, the fungal colonies were counted, recorded and the Colony of Forming Units (CFU) per gram were calculated. Identification was performed by cultural and morphological characteristics and followed taxonomic schemes for Aspergillus and Penicillium (Samson and Pitt, 2000).

Evaluation of Bacterial Contamination
Every sample was examined for bacterial contamination. 0.2 g of each sample was placed on blood agar plates and incubated for 24 h. Bacterial growth was identified using the API method. Pure colonies were isolated and transferred to blood agar plates for antibiotic susceptibility testing.

Table 1: Herbal plants analysed
Image for - Microbial and Heavy Metals Contamination of Herbal Medicines
*http://www.wikipedia.org/wiki/list_of_plants_used_as_medicine
http://www.impgc.com/index.php (Indian MedicinalPlants Growers` Consortium

Evaluation of Toxic Heavy Metal Contamination
All 32 samples were analyzed for toxic heavy metal contamination. Microwave-assisted acid digestion was used for all of the samples using Multiwave 3000 Platform system (Anton Paar, Graz, Austria) and the elemental contents and their infusions were determined by FAAS and ICP-AES. Microwave digestion procedure was applied under optimized conditions for dissolution of herbal plants.

Microwave Acid Digestion and FAAS
In 400 mL cylinder 300 mL concentrated HCl and 100 mL of concentrated HNO3 were added. Mixture was transferred to 1000 mL volumetric flask and volume was filled with diH2O. Mixture was inverted to mix and allowed to stand. Samples were dried at 70°C for 24 h after which they were grinded in Spex mill. Crucibles and caps were prepared by washing in 10% HNO3 and muffle at 750°C for 2 h. Ground samples of 0.5 g were placed into each crucible using Mettler (four-deck) balance and placed in muffle furnace to bring to ashing temperature (450°C) slowly for 90 min and ashed for 4 h. Crucibles were allowed to cool. Linear range of concentrations was determined for the wavelength to be used and an appropriate sample dilution scheme was devised.

Diluted extract was measured for metal content by atomic absorption spectrophotometry (Liberty spectrometer ICP-AES Series II, Varian Australia Pvt. Ltd., Mulgrave, Australia). Using FAAS for the determination of metals, the samples are mineralized in order to avoid possible matrix-related interferences. Iron, copper, zinc, manganese, nickel, cobalt, cadmium and lead content was determined by analyzing the sample solutions undiluted due to low concentration of these metals. Cadmium and lead are complexed by ammonium pyrrolidine dithiocarbamate (APDC) and extracted into methyl isobutyl ketone before they were measured. Ionization of calcium and magnesium was avoided by adding NaCl solution to sample solutions. Chemical interference form phosphates and other anions in measurements of Ca, Mg, Fe, Cu, Zn and Mn are minimized by adding La(NO3)3 solutions to sample solutions.

RESULTS

Table 2 shows the total fungal count, total plate count, fecal coliform count and specific bacterial counts in 32 herbal plants analyzed. Of the 32 herbs analyzed, Astralagus sarcocolla had the highest TPC count (12x105 cfu g-1) followed by Pimpinella anisum (16x104 cfu g-1), Matricaria chamomilia (1x104 cfu g-1), Carum carvi (76x103 cfu g-1) and Nigella sativa (46x103 cfu g-1). Matricaria chamomilia had the highest total coliform count (1x104 cfu g-1) followed by mixed grained herbs (32x103 cfu g-1), Astralagus sarcocolla (30x103 cfu g-1) and Ducrosia ismaelis (29x103 cfu g-1). Matricaria chamomilia also had the highest fecal coliform count (70x104 cfu g-1) followed by Astralagus sarcocolla (15x103 cfu g-1), mixed grained herbs (70x102 cfu g-1), Ducrosia ismaelis (12x102 cfu g-1) and mixed herbs (10x102 cfu g-1). Zingiber officinale had the highest Staphylococcal content with 77x103 cfu g-1, followed by grained mixed herbs (37x103 cfu g-1) and Cinnamomum zeylanicum (35x103 cfu g-1). Grained mixed herbs had the highest Bacillus cereus content with 10x103 cfu g-1, followed by Astralagus sarcocolla (2x103 cfu g-1), Anastatica hierochuntica (17x102 cfu g-1) and Zingiber officinale (5x102 cfu g-1). Aspergillus flavus and Aspergillus fumigatus dominated the picture in twenty-one (60%) of the analyzed herbal plants with 9 isolates of 35 (25.7%) for each. Whereas Aspergillus ochraceus was isolated from 8 of 35 samples (22.9%).

Table 2: Total fungal count, total plate count, fecal coliform count and specific bacterial counts in 32 herbal plants analyzed
Image for - Microbial and Heavy Metals Contamination of Herbal Medicines
TPC: Total (bacterial) Plate Count, TCC: Total Coliform Count, FCC: Fecal Coliform Count, TFC: Total Fungal Count, cfu: Colony forming units

Table 3: Heavy metal contents (in ppm) of 32 herbal plants analyzed
Image for - Microbial and Heavy Metals Contamination of Herbal Medicines
*values are expressed as ppm of samples analyzed

Table 3 shows the heavy metal contents of 32 herbal plants analyzed. Two henna samples showed a lead content of more than 1 ppm (1.528 and 1.214 ppm, respectively). The rest of the samples showed a lead content of less than 1.0 ppm. Mercury content was the highest in Lepidium sativum with 0.630 ppm of lead, followed by Artemisia herba alba (0.102 ppm), henna (0.092 ppm), Pimpinella anisum and mixed herbs (both with 0.087 ppm each). The rest of the samples had mercury content less than 0.08 ppm. Higher contents of aluminum were recorded in Zingiber officinale (19.83 ppm) followed by Anastatica hierochuntica (17.95 ppm) and Pimpinella anisum (14.28 ppm) the rest of the samples had aluminum content less than 10 ppm. Calcium content was the highest in Artemisia herba alba (1.319 ppm) followed by Aloe vera (1.287 ppm), Trigonella foenum (1.281 ppm), Zingiber officinale (1.179 ppm) and Astralagus sarcocolla (1.164 ppm), Boswelia carterii (1.162 ppm), Anethum graveolens (1.288 ppm), Lepidium sativum (1.069 ppm) and Cuminum cyminum (1.055 ppm). The rest of the samples had calcium content less than 1.0 ppm. Cadmium content was highest with Lepidium sativum, Vigna radiata and Zingiber officinale (all had 0.025 ppm of cadmium). The rest of the samples had cadmium content less than 0.02 ppm. Copper was highest with Cinnamomum zeylanicum (0.284 ppm) followed by Matricaria chamomilia (0.282 ppm), Carum carvi (0.274 ppm), Nigella sativa (0.271 ppm), Foeniculum vulgare (0.267 ppm), Achillea fragrantissima (0.244 ppm) and Commiphora myrrah (0.207 ppm). The rest of the samples had copper content less than 0.2 ppm. Iron was highest in Zingiber officinale with 19.44 ppm followed by Matricaria chamomilia (18.56 ppm), Anastatica hierochuntica (16.72 ppm) and Salvia officinalis (13.25 ppm). The rest of the samples had iron content less than 10 ppm. Zinc was the highest in Salvia officinalis (0.962 ppm) followed by Nigella sativa (0.956 ppm), Lepidium sativum (0.877 ppm) and Cinnamomum zeylanicum (0.817 ppm). The rest of the samples had zinc content less than 0.8 ppm. Potassium was the highest in Matricaria chamomilia (943.4 ppm) followed by Zingiber officinale (635.1 ppm), Coffea arabica (555.4 ppm), Cuminum cyminum (515.7 ppm) and mixed herbs (500.8 ppm). The rest of the samples had potassium content less than 500 ppm. Pimpinella anisum showed the highest content of sodium (307.4 ppm) followed by Matricaria chamomilia (227.7 ppm), Ducrosia ismaelis (155.4 ppm), henna (102.1 ppm) and Foeniculum vulgare (100.5 ppm). The rest of the samples had sodium content less than 100 ppm.

Microbial analysis of 32 herbal plants showed that Bacillus species was seen in 3 (9.7%) of the isolated microorganisms of which Bacillus cereus dominated with 14 isolates (45.2%). Other microbial isolates were Aeromonas hydrophilia, Shigella spp., Enterobacter agglomerans, Enterobacter spp., Vibrio fluvialis, Escherichia coli, Pasteurella multocida, Enterobacter cloacae, Staphylococcus hyicus, Staphylococcusepidermidis, Acinetobacter iwoffii and Klebsiella (Table 4). Sensitivity testing for antibiotics showed that most of these isolated microorganisms were sensitive to amoxicillin, gentamicin, imipinem, tobramycin and trimethoprim-sulfamethoxazole. Enterobacter cloacae showed resistance to ampicillin and cefazolin, whereas Aeromonas hydrophilia showed resistance to cefotaxime and ceftazidime. Shigella spp. showed resistance to cefazolin and Escherichia coli showed resistance to ciprofloxacin (Table 5).

Table 4: Microbial isolates from 31 herbal plants analyzed
Image for - Microbial and Heavy Metals Contamination of Herbal Medicines

Table 5: Bacterial sensitivity test results of microbial isolates from analyzed herbal plants
Image for - Microbial and Heavy Metals Contamination of Herbal Medicines
S = Sensitive, R = Resistant, Amox: Amoxicillin, Ampi: Ampicillin, Cefaz: Cefazolin, Cefta: Ceftazidime, Cipro: Ciprofloxacin, Genta: Gentamicin, Imipi: Imipinem, Tobra: Tobramycin, Trim: Trimethoprim/sulfamethoxazole

DISCUSSION

In our study, Aspergillus spp. was the predominant fungi recovered and as such the major toxigenic species. Twenty-one (65.6%) of our samples analyzed had Aspergillus flavus and Aspergillus fumigatus dominating the picture. Five (15.6%) had enumeration limits of more than 2x102, the enumeration limit for total fungal count as set by the US Pharmacopoeia. This high percentage of fungal isolation form herbal plants as revealed by our results may indicate the inherent capacity of these moulds to instigate deleterious effects on humans when consumed. Aside from these, microorganisms have been isolated from our samples with Bacillus cereus isolated in 14/31(45.2%) of samples. Bacillus cereus is an endemic, soil dwelling bacteria that causes food borne illness (Ryan and Ray, 2004). When ingested, this microbe causes severe nausea, vomiting and diarrhea. Generally speaking, Bacillus foodborne illnesses occur due to survival of the bacterial spores when food is improperly cooked. Escherichia coli can also cause serious food poisoning in humans. In fact, this organism has been used widely as an indicator of fecal contamination in water. Shigellla cause dysentery by invading the colonic mucosa. They cause cell death and spread laterally causing mucosal ulceration, inflammation and bleeding. Severe infection with Shigella and its subspecies include hemolytic-uremic syndrome, seizures, sepsis and toxic megacolon in humans. Vibrio fluvialis has been associated with diarrhea although they have been rarely isolated (Hickman-Brenner et al., 1984). Acinetobacter iwoffii can cause severe respiratory disease (Robino et al., 2005). Pasteurella multocida has been associated with zoonotic infection in humans and Klebsiella with chronic diseases of the upper airways (Bothello-Nevers et al., 2006). Enterobacter agglomerans and Staphylococcus epidermidis are generally prevalent in the environment and usually relatively benign, it does have a potential for nosocomial infection (Geere, 1977). Furthermore, the sensitivity testing results in this study showed that most of the organisms isolated are sensitive to the more common and available antibiotics, thus, available treatment protocols are in place for such bacterial colonization in humans.

Present study also showed contamination with heavy metals, some of them toxic to humans. In general, plants do not absorb or accumulate lead however, in soils with high lead content; it is possible for some lead to be taken up. Soils with lead levels exceeding 100 ppm should not be used for gardening and plants can be safely eaten from soils with soil lead levels up to 300 ppm for leafy and root type vegetables (http://www.grayenvironmental.com). Limit of quantification for lead in herbal medicine should not exceed 2.0 ppm. Lead was found in all of our samples, of which 2 henna samples showed 1.2-1.5 ppm lead content. This level however was not considerably high to alarm for human use or consumption. Similarly, trace levels of mercury and cadmium were present in our samples of up to 0.1 ppm. Limit of quantification for mercury is up to 0.5 and 0.20 ppm for cadmium. The highest recorded amount of mercury was 0.102 in Artemisia herba alba Whereas the highest level of cadmium was 0.025 ppm recorded in 3 samples. These levels of mercury and cadmium do not appear to be of health concern. Aluminum level was high in three herbs (Anastatica hierochuntica, Salvia officinalis and Zingiber officinale) of levels 14.28-19.83 ppm. This is way above the maximum allowed level of 0.2 ppm. Aluminum, as a metal when present in our food, water supply and soil can induce individuals to suffer from aluminum toxicity. After years of accumulated exposure and storage in our body, it can result to brain degeneration and skeletal deformities (http://www.drpepi.com). It is believed that Alzheimer`s disease is related to aluminum toxicity (Derouesne, 2004).

Minerals such as magnesium and zinc are just as critical to maintaining optimal health, or that, taken in excess, these minerals can be toxic. Iron is essential for blood cells; potassium is needed for a healthy nervous system and zinc to enhance immunity and for reproductive function. However, when taken in amounts over the recommended maximum allowable range, they can be toxic to health. These effects occur in nervous system. However, our study showed that none of our samples exceeded the maximum allowable level of 5 ppm for zinc. Although 4 of samples had iron levels more than the maximum allowed level of 15 ppm, these levels were not significantly high. Similarly, most of our samples have high levels of potassium and sodium. Although potassium and sodium are essential to health, excess of potassium can cause cardiac dysfunction and excess sodium can cause metabolic problems and hypertension.

The results of this study especially on the heavy metal contents of herbal plants implicate an impending danger for consumers. Significant serious consequences may appear due to accumulation of heavy metal contents through years of frequent use of these herbal medicines. Several degenerative and life-threatening conditions were linked to accumulation of toxic metals in the body. When our body is compounded with microbial infection to an already toxin-filled system, the capacity of our immune system to defend itself is exhausted. Although this study showed heavy metal levels within the allowable limits, it is possible that some amounts can be taken up by the system and accumulate for years of use, thus cause serious consequences. Even if these metals found in herbal medicines are less likely than free to bind with molecules in our body and thus slower to be absorbed, the issue of safety and vigilance on its serious adverse effects be of a concern. Furthermore, the continued practice of safety and precautionary measures from the harvest area (e.g., minimization of pesticide use) to the household (e.g., thorough cleaning and washing of herbal plants prior to use) should be practiced.

CONCLUSION

There are considerable amounts of microbial contamination and heavy metal concentrations in commonly used herbal plants. Consumers have the right to be informed and must be warned about health hazards through proper signs, labels and brochures indicating possible dangers lurking in their food and household products.

REFERENCES

  1. Ang, H.H., 2003. Analysis of lead content in herbal preparations in Malaysia. Hum. Exp. Toxicol., 22: 445-451.
    CrossRef  |  


  2. Ang, H.H. and K.L. Lee, 2006. Contamination of mercury in tongkat Ali hitam herbal preparations. Food Chem. Toxicol., 44: 1245-1250.
    CrossRef  |  Direct Link  |  


  3. Botelho-Nevers, E., F. Gouriet and H. Lepidi, 2006. Chronic nasal infection caused by Klebsiella rhinoscleromatis or Klebsiella ozaenae: Two forgotten infectious diseases. Int. J. Infectious Dis., 11: 423-429.
    CrossRef  |  PubMed  |  Direct Link  |  


  4. Boyd, N.D., H. Benediktsson, M.J. Vimy, D.E. Hooper and F.L. Lorscheider, 1991. Mercury from dental silver tooth fillings impairs sheep kidney function. Am. J. Physiol., 30: 1010-1014.
    PubMed  |  


  5. Caldas, E.D. and L.L. Machado, 2004. Cadmium, mercury and lead in medicinal herbs in Brazil. Food Chem. Toxicol., 42: 599-603.
    CrossRef  |  PubMed  |  Direct Link  |  


  6. Cornett, C.R., W.R. Markesbery and W.D. Ehmann, 1998. Imbalances of trace elements related to oxidative damage in Alzheimers disease brain. Neurotoxicity, 19: 339-346.
    PubMed  |  


  7. Derouesne, C., 2004. The role of aluminum in the genesis of Alzheimer’s disease relaxes in the absence of sufficient proof in the current state of our knowledge. Neurotoxicity of aluminum: Doubt for highly exposed population. Psychol. Neuropsychiatr Vieil., 2: 76-76.
    PubMed  |  


  8. Echeverria, D., H.V. Aposhian, J.S. Woods, N.J. Heyer and M.M. Aposhian et al., 1998. Neurobehavioral effects from exposure to dental amalgam Hg: New distinctions between recent exposure and Hg body burden. FASEB J., 12: 971-980.
    Direct Link  |  


  9. Frustaci, A., N. Magnavita, C. Chimenti, M. Caldarulo and E. Sabbioni et al., 1999. Marked elevation of myocardial trace elements in idiopathic dilated cardiomyopathy compared with secondary cardiac dysfunction. J. Am. Coll. Cardiol., 33: 1578-1583.
    Direct Link  |  


  10. Geere, I.W., 1977. Enterobacter agglomerans: The clinically important plant pathogen. Can. Med. Assoc. J., 116: 517-519.
    Direct Link  |  


  11. Han, C., J. Li and Q. Hui, 2008. Determination of trace elements in Jinqi a traditional Chinese medicine. Biol. Trace Elem. Res., 122: 122-126.
    CrossRef  |  PubMed  |  


  12. Hickman-Brenner, F.W., D.J. Brenner, A.G. Stegerwalt, M. Schreiber and S.D. Holmberg et al., 1984. Vibrio fluvialis and Vibrio furnissii isolated from a stool sample of one patient. J. Clin. Microbiol., 20: 125-127.
    Direct Link  |  


  13. Ichinoe, M., H. Konuma, A. Kartastina and M. Satake, 1998. Microbial contamination of traditional herbal drugs in Indonesia. Eisei Shikenjo Hokoku, 106: 18-24.
    PubMed  |  


  14. Kneifel, W., E. Czech and B. Kopp, 2002. Microbial contamination of medicinal plants-a review. Planta Med., 68: 5-15.
    PubMed  |  


  15. Ngim, C., 1989. Epidemiologic study on the association between body burden mercury level and idiopathic parkinson’s disease. Neuroepidemiology, 8: 128-141.
    CrossRef  |  


  16. Nordic Committee on Food Analysis (NMKL), 1999. Coagulase Positive Staphylococci. Enumeration in Foods NMKL Method No. 66. 3rd Edn., Nordic Committee on Food Analysis, Oslo, Norway
    Direct Link  |  


  17. Nordic Committee on Food Analysis (NMKL), 2003. Bacillus Cereus. Determination in Foods, NMKL Method No. 67. 4th Edn., Nordic Committee on Food Analysis, Oslo, Norway


  18. Nordic Committee on Food Analysis (NMKL), 2004. Coliform Bacteria. Determination in Foods and Feed, NMKL Method No. 44. 5th Edn., Nordic Committee on Food Analysis, Oslo, Norway
    Direct Link  |  


  19. Nordic Committee on Food Analysis (NMKL), 2005. Thermotolerant Coliform Bacteria and E. coli Enumeration in Food and Feed, NMKL No. 125. 5th Edn., Nordic Committee on Food Analysis, Oslo, Norway
    Direct Link  |  


  20. Robino, P., E. Bert, C. Tramuta, S.S. Ceruti and P. Nebbia, 2005. Isolation of acinetobacter iwoffii from a lovebird with severe respiratory symptoms. Schwizer Arch. fur Tierheilkunde, 147: 267-269.


  21. Ryan, K.J., C.G. Ray and J.C. Sherris, 2004. Sherris Medical Microbiology: An Introduction to Infectious Diseases. 4th Edn., McGraw Hill, New York, USA., ISBN-13: 9780838585290, Pages: 979


  22. Vimy, M.J., Y. Takahashi and F.L. Lorscheider, 1990. Maternal-fetal distribution of mercury released from dental amalagams. Am. J. Physiol., 27: 939-945.
    Direct Link  |  


  23. Yasue, H., T. Itoh, T. Mizuno Y and E. Harada, 2007. Severe hypokalemia, rhabdomyolysis, muscle paralysis and respiratory impairment in a hypertensive patient taking herbal medicines containing licorice. Int. Med., 46: 575-578.
    CrossRef  |  PubMed  |  


  24. Samson, R.A. and J.I. Pitt, 2000. Integration of Modern Taxonomic Methods for Penicillium and Aspergillus Classification. 1st Edn., Harwood Academic Publishers, Amsterdam, pp: 510


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