Parasites, microbial contaminants, mycotoxins and endotoxins are the classes of biological contaminants likely to be found in herbal materials.

1. Parasites: Protozoa, nematodes and their ova may be introduced during cultivation or during processing and manufacturing if personal hygiene measures have not been taken by handling personnel. As a result zoonosis may occur especially if uncomposted animal excreta are used during cultivation.
2. Microbiological contaminants: Medicinal plants may be associated with a broad variety of microbial contaminants represented by bacteria, fungi and viruses. This microbiological background depends on several environmental factors and affects the overall quality of herbal products and preparations.

A large number of bacteria and moulds often originating in soil or derived from manure are found in herbal materials. Some of them form the naturally occurring microflora of medicinal plants of which aerobic spore-forming bacteria frequently predominate. Improper methods of harvesting, cleaning, drying, handling and storage may cause additional contamination and microbial growth.

Microbial contamination may also occur during processing of herbal materials. Failure to control moisture levels during transportation and storage as well as failure to control the temperature of liquid formulations and finished products could be the reason. Presence of Escherichia coli and Salmonella species and moulds may indicate poor quality of production and harvesting practices. GAP and GMP are thus critical for effective control of microbial contamination. Laboratory procedures investigating microbial contamination limit their levels as indicated by values laid down in pharmacopoeias as well as in WHO guidelines. Generally a complete procedure consists of determining the total aerobic microbial count, the total fungal count and the total Enterobacteriaceae count, together with tests for the presence of E. coli, Staphylococcus aureus, Shigella, Pseudomonas aeruginosa and Salmonella species. As per WHO guidelines, Salmonella and Shigella species must not be present at any stage in herbal medicines intended for internal use. Different pharmacopoeias have different testing requirements.

Vegetable drugs tend to show much higher levels of microbial contamination than synthetic products. European pharmacopoeias hence allow higher levels of microbial contamination in herbal drugs than in synthetic pharmaceuticals. The allowed contamination level may also depend on the method of processing of the drug. For example, higher levels are permitted if the final herbal preparation involves boiling with water. Presence of fungi should be carefully monitored since common species produce toxins which can pose acute and chronic risks to health.

These fungal toxins called mycotoxins are usually secondary metabolic products which are nonvolatile, have a relatively low molecular weight and may be secreted onto or into the medicinal plant material. They may help parasitic fungi invade host tissues. Mycotoxins are of four main groups namely aflatoxins, ochratoxins, fumonisins and trichothecenes, all of which have toxic effects.

Aflatoxins have been extensively studied and are classified as group I human carcinogens by the international agency for research on cancer. Aflatoxin-producing fungi sometimes build up during storage. Procedures for their determination in herbal drugs are published by the WHO. After a thorough clean-up procedure, TLC is used for confirmation. Mycotoxins produced by the species of fungi including Aspergillus, Fusarium and Penicillium are the most commonly reported. In addition to the risk of bacterial and viral contamination, herbal materials may also carry bacterial endotoxins. Found mainly in the outer membranes of certain Gram-negative bacteria and released during cell disruption, they are complex lipopolysaccharide molecules that elicit an antigenic response, cause altered resistance to bacterial infections and have other serious effects. Certain plant constituents are susceptible to chemical transformation by contaminating microorganisms. Withering leads to enhanced enzymatic activity, transforming some of the constituents to other metabolites not initially found in the herb. These newly formed constituents may cause adverse effects along with formed moulds such as Penicillium nigricans and P.jensi.

Tests for the presence of microbial contamination on herbal drugs should be performed in dosage forms in compliance with the requirements of national, regional or international pharmacopoeias. BP for example requires drugs such as acacia, agar and powdered digitalis to be free of E. coli in the quantity of the material stated. Others like alginic acid, cochineal and tragacanth are also tested for the absence of Salmonella. Drugs such as agar and guar gum galactomannan should not have total viable count of 10 microorganisms/gm. Generally manufacturers will ensure that for crude drugs to be taken internally, the limits for bacterial and mould contamination as applicable to food stuffs are adhered to. Considerable quantities of drugs are sterilized in special equipment by treatment with ethylene oxide.