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Eleven stored product beetles were examined for surface features of the beetle elytron, using a scanning electron microscope. The 3 characteristics, sculpturing pattern, setae, and setal pits, were well defined and could be used to distinguish the species examined. The scanning electron microscope can give 3-dimensional images at magnifications up to 20,000 ×, in contrast with the light microscope which has a maximum magnification of 1000 ×.

Eleven bleaching agents were used to decolorize cocoa tissues, and their effects on insect fragments and rodent hairs were studied. Gilorine bleaches, potassium permanganate, and hydrogen peroxide all result in good bleaching of ground cocoa. However, chlorine bleaches completely destroy rodent hair, and potassium permanganate is not effective for high fat products. Hydrogen peroxide decolorizes cocoa bean tissue very well and does not destroy rodent hairs nor decolorize insect fragments. A mechanical means of evaluating the amount of bleaching of cocoa bean tissues is also described. Future studies are planned on the use of hydrogen peroxide as a plant tissue bleaching agent

A staining method is described in which plant material is differentially stained from insect fragments, rodent hairs, and bird feather barbules. Because the colors of the stained plant tissue contrast with that of the dyed extraneous material, counting of the contaminants becomes easier, rapid, and more accurate.

A new method has been developed which is applicable to all nutmeats, with or without testa, either raw, dry, or oil roasted. Nutmeats are defatted by a 10 min boiling in chloroform (pecans require 3 defattings), after which the solvent is removed by aspiration and an isopropanol rinse. Following a short boiling deaeration and acid hydrolysis, light filth elements are extracted with mineral oil from a mixture of 60% ethanol and calcium chloride. Collaborative trials by the proposed method for nutmeats (except pecans) yielded significantly cleaner extraction papers and better recoveries of all light filth spike elements than by the official method. In addition, the proposed method resulted in reductions of coefficients of variation ranging from 47 to 93% of that for the official method. The modification for pecans, included in this method, required more extensive defatting and also resulted in improved light filth recoveries and reductions in coefficients of variation. Not all varieties of nutmeats were collaboratively studied; however, intralaboratory recovery trials on these nutmeats proved the proposed method superior to the official methods. The method is recommended for adoption as official first.

The second paper in this series describes some useful and time-saving microanalytical techniques. Topics discussed include sealing of glass bottles, vial closures, and petri dishes, collaborative evaluation forms, permanent identification of metal and glass sample containers, shipping containers for breakable collaborative sample containers, new defatting procedure prior to light filth analysis, new filtration procedures, collection of live mites and insects from bulk food samples, and preparation and handling of microscopic exhibits of insect fragments and hairs.

A method has been developed for the extraction of light filth from seeded and seedless raisins. Raisins are chloroform defatted, rehydrated in water, wet sieved, and extracted in a trap flask. Collaborative study resulted in reproducible recoveries and clean filter papers. This method is recommended for adoption as official first action.

A new method has been developed for the extraction of extraneous materials from ground cinnamon. In the method, hydrochloric acid is used to disperse gel formations, wet sieving is used to eliminate the dispersed gel, and a percolator replaces a trap flask to separate the extraneous material from cinnamon tissue. Collaborative results were good, and the method is recommended for adoption as official, first action.

A method developed for the extraction of light filth from raw and processed wheat germ utilizes chloroform defatting and acid hydrolysis followed by wet sieving. Light filth is extracted with mineral oil from a hot ethanol or isopropanol solution. The collaborative study resulted in 90, 89, and 84% average recoveries of spike insect fragments, rodent hairs, and adult insects, respectively. The method is recommended for adoption as official first action.

Collaborative results are presented on a proposed method for light filth extraction from corn- and rice-based ready-to-eat breakfast cereals and corn-based snack foods. Samples are digested with hydrochloric acid, followed by wet sieving on a No. 230 mesh plain weave sieve. Sieve retainings are boiled in 40% isopropanol and filth is extracted in either a Kilborn funnel or a trap flask. Average per cent recoveries of spiked elytral fragments and rodent hairs were 91.0 and 80.0% (using trap flask) and 86.1 and 81.7 (using Kilborn funnel), respectively. The method is recommended for adoption as official first action.

A rapid extraction method has been developed for the separation of light filth from paprika; in the method, paprika is defatted with a surfactant, wet sieved, and floated with n-heptane from 60 % alcohol and Tween 80-Versene mixture. Collaborative results by the method gave 8.7 and 16.1% increased recovery of insect fragments and rodent hairs, respectively. The method is recommended for adoption as official, first action.

A study is described in which re-heptane is compared with gasoline as flotation oils from water and alcohol lower phases. n-heptane, containing less than 4% toluene, gave higher and more reproducible insect fragment and rodent hair recoveries than gasoline when used with an alcohol lower phase. Recoveries were comparable between n-heptane and gasoline when used with a water lower phase. In both cases filter papers were cleaner or contained equal amounts of plant debris as obtained with gasoline. It is recommended that re-heptane replace gasoline for use as the flotation oil in alcohol and water extractions, and that "n-heptane# replace the word "gasoline# in method 36.002(o).

Insect chewing on food packaging materials has been characterized in order to differentiate between entrance and exit bore holes. The characteristics specific to various types of packaging are presented as a method in the form of a descriptive “key” that permits determination of the direction of boring. A collaborative study of the method resulted in 92% correct determinations of boring direction. The method has been adopted as official first action.

A method for the extraction of light filth from crude and refined papain has been developed. The product is solubilized by boiling in water, followed by wet sieving. Depending upon the amount of residue remaining on the sieve, the product is either transferred directly to filter paper(s) or trapped off in a Wildman trap flask. Collaborative recoveries of spiked insect fragments and rodent hairs were 94 and 92%, respectively. The method has been adopted as official first action.

The official AOAC method for filth in starches, 40.056, has been modified to obtain better recovery of extraneous materials. In lieu of wet sieving on a No. 140 sieve, the modified method utilizes No. 230 wet sieving followed by an acid hydrolysis, if excessive starch material remains on the sieve. A limited collaborative study comparing the improved method with the official method resulted in higher recoveries of rodent hairs and equivalent recoveries of other spike elements by the improved method. The improved method is recommended for adoption as official first action to replace 40.056.

A rapid procedure for the isolation of curculio larvae from pecan pieces has been studied collaboratively. The larvae are separated from 115 g portions of nutmeats by notation from isopropanol. The collaborative study showed that the method requires approximately 10 min analytical time/1 lb sample; average recovery of curculio larvae was 99%. The method is recommended for adoption as official first action.

Collaborative results are presented on a new method for filth in casein and sodium caseinate. The method consists of dispersing the product in a Versene solution, wet sieving on a No. 230 sieve, and transferring filth directly from sieve onto filter paper. The collaborative study resulted in an average recovery of 98 and 96% for insect fragments and rodent hairs, respectively. The method is recommended for adoption as official first action.

A new method has been developed for the extraction of extraneous materials from plant gums. Hydrochloric acid and autoclaving are used to break down and disperse partially insoluble gums, followed by sieving and extracting using a trap flask. Collaborative studies gave reproducible recoveries of rodent hairs and insect fragments, and the method is recommended for adoption as official first action.

In the method described, the black or white ground pepper sample is slurried with water and wet sieved. The pepper residue is defatted during the sieving step. Flotation extraction of the sieved material utilizes a combination of n-heptane-mineral oil as the oil layer over 40% isopropanol. Collaborative studies resulted in reproducible recoveries and light-to-medium laden filter papers. It is recommended that the method be adopted as official first action to replace 40.109(b).

In the method described for the extraction of light filth from wheat gluten, the product is dispersed in an alcohol-acid solution and then boiled, with stirring. After boiling, the product is trapped off with mineral oil from an aqueous solution of 40% isopropanol in a Wildman trap flask. In the collaborative study reproducible counts and high recoveries of insect fragments and rodent hairs were obtained and the method was demonstrated to be rapid and easy to perform. The method is recommended for adoption as official first action.

A method has been developed for the extraction of light filth from wheat-based ready-to-eat breakfast cereals. The method utilizes acid digestion followed by wet sieving and washing with isopropanol, then trapping with a heptane-paraffin oil mixture from a sodium oleate-40% iso propanol solution. A collaborative study resulted in 75% average recoveries of spike elytral squares and rodent hairs from wheat-based cereals. The method, as developed for oat-based cereals, resulted in debris-laden filter papers and therefore was judged ineffective. The method has been adopted as official first action for wheat-based cereals. Further studies are recommended for oat-based cereals.

A method has been developed for the extraction of light filth from prepared horseradish. The method involves a direct flotation from 40% isopropanol with a mineral oil-n-heptane mixture. Collaborative study showed the method to be rapid and easy to perform, and to yield good recoveries and clean extraction papers. The method has been adopted as official first action.

A new method has been developed for the extraction of extraneous materials from unground cassia. Hydrochloric acid is used to remove adhering extraneous material from the bark; double sieving is used to separate larger pieces of bark from filth elements and smaller plant material; and extraction of extraneous material from plant debris is performed in a trap flask. Collaborative results were good, and the method is recommended for adoption as official first action.

The bleaching agent used for the extraction of light filth from cocoa, chocolate, and press cake has been changed from hydrogen peroxide to sodium hypochlorite. Although hydrogen peroxide has many advantages, it also bleaches light colored fragments of insect cuticles, thus making counting difficult. The change has been adopted as official and 44.006(d) will be revised.

Ground cocoa bean tissue is decolorized with hydrogen peroxide, then stained with picric acid to make decolorized hairs visible. Hydrogen peroxide does not destroy rodent hairs or decolorize insect fragments as does sodium hypochlorite which has previously been recommended by several researchers as a plant bleaching agent.

A rapid alternative method has been developed for dispersing cocoa press cake in preparation for light fdth analysis. The press cake is broken into pieces one-half inch or smaller and mechanically stirred in hot (150°C) peanut oil for dispersion prior to defatting with Triton X-114. Light filth is then determined by method 40.006(a). Intralaboratory studies proved the method to be faster than the existing dispersion procedures and to result in almost twice the recovery of rodent hairs. Recoveries of insect fragments were not affected. The method has been adopted as an alternative to dispersion techniques in 40.006(c).