Determination of Extraneous Material in Bakery Products Without Fruit and Nut Ingredients

Laboratory Procedure ExFLP-22
April 1997

1. Application

This method is applicable to the sampling and examination of bakery products without fruit and nut ingredients for extraneous material, such as insect infestation, and rodent hairs to determine compliance with Sections 4, 5 and 7 of the Food and Drugs Act. This method replaces ExFLP-22 dated September 1987.

2. Definition of Terms

A lot is defined as that amount (volume, weight, etc.) of the food which is produced, stored and/or shipped under conditions as nearly uniform as possible, preferably designated by a common container code or marking, and, in any event, consisting of not more than one variety, grade or type of product from a single identifiable source.

3. Collection of Samples

• 3.1 Scrutinize the entire lot for live infestation. If live infestation is found, do not sample until after fumigation or other effective treatment.
• 3.2 Obtain three sample units selected at random from the lot of at least 100 g each using appropriate sampling equipment and containers. Three sample units constitute a sample.
• 3.3 Each sample unit must be kept separate and labelled 1, 2 and 3. Complete information respecting the lot size, weight of individual containers, country of origin, exporter, importer, or domestic manufacturer, and product and lot identification should be recorded and should accompany the sample.
  
  

4. Materials and Special Equipment

1. Balance
2. 2-2.0 L beakers with covers (watch glasses)
3. Autoclave. Either (a) slow exhaust type (Set slow exhaust to lower pressure from 1.15 to 0 kg per sq. cm. in 15-20 min.) or (b) non-slow exhaust (Let cool to 0 kg per sq. cm. before opening or venting). Alternatively, instead of an autoclave use a steam bath.
4. Smooth magnetic stirring bar, teflon coated (1x5 cm)
5. Stirrer hot plates
6. U.S. standard No. 230 sieve
7. 2 L percolator, premarked at 250 and 1700 mL levels
8. Rubber policeman attached to a long glass rod
9. Wash bottles
10. Suction filtration apparatus with Buchner or Hirsch funnel (5-7 cm perforated plate)
11. Ruled filter paper. Filter paper should be larger than funnel plate (7-9 cm)
12. Petri dishes to suit size of filter paper used
13. Stereoscopic microscope (10-30x)
14. Ashless filter paper. Filter paper should be larger than funnel plate (7-9 cm)
15. Drying oven (40-60°C)
16. Crucible
17. Desiccator
18. Muffle oven
19. Emulsifiers, nonionic, water-soluble surfactants.
    
    
    1. Dialkylphenoxypoly (ethyleneoxy) ethanol - Igepal DM-710 (GAF Corporation).
    2. Nonylphenoxypoly (ethyleneoxy) ethanol - Igepal CO-730 (GAF Corporation).
20. Concentrated HCl
21. Antifoam solution. Antifoam "B" (BDH Chemicals)
22. Mineral oil. Paraffin oil, white, light 125/135 Saybolt Universal Viscosity (38°C), specific gravity 0.840-0.860 (24°C)
23. Alcohol, either 95% ethanol or 99% isopropanol
24. Detergent (1% sodium lauryl sulphate)
25. Glycerol-alcohol (95%) mixture (1:1)
26. Chloroform (CHCl₃)

5. Procedure

The examination shall be carried out in accordance with the following instructions.

• 5.1 Preparation of Analytical Units
  • 5.1.1 Thoroughly mix a sample unit and in a random fashion weigh 100 g. This 100 g constitutes an analytical unit.
  • 5.1.2 Repeat Step 5.1.1 for each of the remaining two sample units.
• 5.2 Isolation - Light Filth
  
  
  • 5.2.1 Add 1 L hot (55-70°C) tap water to a 2 L beaker. Add 20 mL of emulsifier Igepal DM-710 and 5 mL of emulsifier Igepal CO-730, and mix well. Add a 100 g analytical unit, breaking into pieces < 4 sq. cm. Stir well. Proceed with digestion (a) or (b).
    
    (a) Autoclave. Add 30 mL of HCl with stirring and 1 mL of antifoam solution. Autoclave for 30 min. at 121°C.
    
    (b) Steam bath. Add 90 mL of HCl with stirring. Heat in a steam bath for 10 min. Add 1 mL of antifoam solution. Boil for 15 min. on a magnetic stirrer-hot plate, keeping the beaker covered with a watch glass.
  • 5.2.2 Transfer slurry to a No. 230 sieve. Wash contents of sieve with a stream of hot (50-70°C) tap water until washwater runs clear and the water runs freely through the material.
  • 5.2.3 Transfer sieve retainings to original beaker with hot water (50-70°C) and dilute to approximately 1 L. Add 30 mL of HCl and a magnetic stirring bar. Bring to a boil and boil 6 min.
  • 5.2.4 Add 50 mL of mineral oil and continue heating until boiling resumes. Transfer beaker to a cool magnetic stirrer plate and stir magnetically at maximum speed that does not produce visible or audible splashing for 3 min.
  • 5.2.5 Promptly transfer contents of beaker to a percolator containing approximately 250 mL of water. Rinse contents of beaker into percolator with hot tap water making certain to transfer all material including heavy filth. Bring volume in percolator to 1700 mL with hot tap water. After 1 min., stir percolator contents with a long glass rod.
  • 5.2.6 Using wash bottles, rinse magnetic particles from stirring bar into a beaker first with water, then alcohol and finally with detergent until all particles are washed off. Filter washings through ruled filter paper using suction filtration apparatus. Transfer filter paper to a petri dish and moisten with glycerol-alcohol mixture.
  • 5.2.7 After 2 min., drain oil interface in percolator to 250 mL into a 2 L beaker. Retain contents for heavy filth determination.
  • 5.2.8 Refill percolator by pouring hot tap water down percolator sides to loosen adhering material and refill to 1700 mL mark.
  • 5.2.9 After 2-3 min., drain percolator to 250 mL, discard drainings and refill for 2 or more cycles. Lower layer should be almost free of suspended material after refill. If not continue through >= 1 recycles.
  • 5.2.10 Finally, drain oil-water interface to 250 mL mark and drain remaining contents into original beaker. Promptly, using wash bottles, wash down sides of percolator successively with >= 50 mL portions of hot tap water, alcohol, hot tap water and/or detergent.
  • 5.2.11 Using suction filtration apparatus, filter beaker contents through ruled filter paper with a minimum of 50 mL washes each of water, alcohol, water and/or detergent. Use a rubber policeman if necessary to clean sides of beaker.
  • 5.2.12 Transfer filter paper(s) to petri dish(es) and moisten with glycerol-alcohol mixture.
  • 5.2.13 Repeat isolation for remaining two analytical units.
• 5.3 Examination - Light Filth
  
  Examine filter papers microscopically at 30x for all types of extraneous material.
• 5.4 Recording Results - Light Filth: ExFLP-22
  • 5.4.1 Classify and record insect fragments into the following two categories:
    
    (a) premilling insect fragments ( <= 0.2 mm)
    
    (b) postmilling insect fragments ( > 0.2 mm)Examples of insects that may infest bakery products are presented in Tables 1-3.
  • 5.4.2 Record the number and size range of all other types of light filth detected. This should be done for each analytical unit separately.
• 5.5 Isolation - Heavy Filth
  • 5.5.1 Stir contents of beaker containing drainings from Light Filth, Step 5.2.7, with a rubber policeman for 30 sec. and allow suspension to stand covered and undisturbed for 30 min.
  • 5.5.2 Decant suspended food material without disturbing bottom contents.
  • 5.5.3 Add 400-500 mL of water, stir 30 sec. and let stand 10 min.
  • 5.5.4 Repeat steps 5.5.2-5.5.3 until practically all plant tissue has been decanted.
  • 5.5.5 If any heavy filth is present, add 100-200 mL of CHCl₃, stir 30 sec., let stand 10 min. and then decant water layer without disturbing or pouring off any of CHCl₃-water interface.
  • 5.5.6 Filter contents of beaker onto an over-dried preweighed ashless paper. Transfer filter paper to petri dish.
• 5.6 Examination - Heavy Filth
  
  Examine filter paper microscopically at 30x for rodent pellets, insect excreta, insect larvae, insect eggs and other organic filth.
• 5.7 Recording Results - Heavy Filth: ExFLP-22
  • 5.7.1 Record the number and size range of each type of heavy filth detected. This should be done for each analytical unit separately.
  • 5.7.2 Dry filter paper in an oven and weigh. Determine weight of heavy filth by difference, and record.
  • 5.7.3 If there is an appreciable amount of residue, transfer filter paper with contents to a preweighed crucible and ash in a muffle oven at approximately 500°C for two hr. Cool material in a desiccator and weigh. Determine ashed weight by difference, and record.
    
    

6. Interpretation

• 6.1 See Table 2 of "Health Protection Branch Standards and Guidelines for Microbiological Safety and General Cleanliness of Food - An Overview" found in Volume 1 of the Compendium of Analytical Methods for HPB guidelines relating to different types of extraneous materials in bakery products without fruit and nut ingredients.
• 6.2 If there is a question as to the acceptability of the lot, contact Evaluation Division, Bureau of Microbial Hazards, Food Directorate, Health Protection Branch, (Phone (613) 957-0349 or FAX (613) 952-6400).
  
  

7. References

• 7.1 Official Methods of Analysis of the Association of Official Analytical Chemists (AOAC) 1990, 15th ed., Arlington, Va. 22201 U.S.A.
• 7.2 Thrasher, J.J. 1970. Acidic mineral oil extraction of light filth from bread and donuts. J. Ass. Off. Anal. Chem. 53:562-566.

Table 1 - Examples of field insects

Common Name	Order
armyworms	Lepidoptera
sawflies	Hymenoptera
jointworms	Hymenoptera
strawworms	Hymenoptera
grasshoppers	Orthoptera
aphids	Homoptera
bugs	Hemiptera
hessian flies	Diptera
midges	Diptera
thrips	Thysanoptera

Table 2 - Examples of insects infesting whole undamaged grains in storage. These insects may also attack grains in the field in warm climates

Species	Family	Common Name
ORDER: Coleoptera
Rhyzopertha dominica (Fabricius)	Bostrichidae	lesser grain borer
Sitophilus granarius (L.)	Curculionidae	granary weevil
Sitophilus oryzae (L.)	Curculionidae	rice weevil
Sitophilus zeamais Motschulsky	Curculionidae	maize weevil
ORDER: Lepidoptera
Sitotroga cerealella (Olivier)	Gelechiidae	Angoumois grain moth

Table 3 - Common insects infesting damaged and milled grain

Species	Family	Common Name
ORDER: Coleoptera
Alphitobius diaperinus (Panzer)	Tenebrionidae	lesser mealworm
Cryptolestes capensis (Waltl)	Cucujidae	Cape grain beetle
Cryptolestes ferrugineus (Stephens)	Cucujidae	rusty grain beetle
Cryptolestes pusillus (Scho"nherr)	Cucujidae	flat grain beetle
Cryptolestes turcicus (Grouvelle)	Cucujidae	Turkish grain beetle
Gnathocerus cornutus (Fabricius)	Tenebrionidae	broadhorned flour beetle
Lasioderma serricorne (Fabricius)	Anobiidae	cigarette beetle
Latheticus oryzae Waterhouse	Tenebrionidae	longheaded flour beetle
Oryzaephilus mercator (Fauvel)	Cucujidae	merchant grain beetle
Oryzaephilus surinamensis (L.)	Cucujidae	sawtoothed grain beetle
Ptinus ocellus Brown	Ptinidae	Australian spider beetle
Stegobium paniceum (L.)	Anobiidae	drugstore beetle
Tenebrio molitor L.	Tenebrionidae	yellow mealworm
Tenebrio obscurus Fabricius	Tenebrionidae	dark mealworm
Tenebroides mauritanicus (L.)	Trogositidae	cadelle
Tribolium castaneum (Herbst)	Tenebrionidae	red flour beetle
Tribolium confusum Jacquelin du Val	Tenebrionidae	confused flour beetle
Trogoderma granarium Everts	Dermestidae	khapra beetle
ORDER: Lepidoptera
Anagasta kuehniella (Zeller)	Pyralidae	Mediterranean flour moth
Cadra cautella (Walker)	Pyralidae	almond moth
Corcyra cephalonica (Stainton)	Pyralidae	rice moth
Ephestia elutella (Hu"bner)	Pyralidae	tobacco moth
Plodia interpunctella (Hu"bner)	Pyralidae	Indianmeal moth
Sitotroga cerealella (Olivier)	Gelechiidae	Angoumois grain moth

NOTE: Insects such as cockroaches and flies that are associated with sewers, excreta and filth in general should not be tolerated.