Detectives discuss digestion differences to deduce the details of fat determination. Don’t dilly-dally; discover the details!

Chapter 42
📂 Case overview: The detectives’ discussion of all the food they ate over the holiday period leads to an in-depth analysis of free and total fat determination. Shallot Holmes takes center stage to improve the detectives’ understanding of a critical process for their clients in the food and feed industry. The importance of hydrolysis in total fat determination is examined along with several issues that could hinder efficiency and reproducibility.

The detectives reconvened after having taken a well-deserved break for the festive season. They sat discussing all the things they had gotten up to over their holidays, which unsurprisingly involved a lot of food. Each of the detectives described in detail all the delicious delicacies they had devoured. Out of all the detectives, Lieutenant Cornlumbo stood out as the detective that had eaten the most. A fact that he was almost too proud of, as demonstrated by the detailed list he had made of everything to have passed his lips on Christmas Day. Not only had he listed the foods, but also the calorific value as if it was some kind of competition. He rolled out the list and went through each item one by one.

“A total of 8,242 calories!” declared Lieutenant Cornlumbo. Miss Mapple looked positively horrified, and Nancy Beef looked as if she might faint. “If you continue to eat like that, your corn will pop,” said Miss Mapple. Cornlumbo said that he was not worried as he had considered the implications. He reminded the detectives about how he had determined their protein content while they were busy working on their Kjeldahl workshops. He said that he had determined his own fat content after Christmas and deduced that he did not contain too much fat, and there was no risk of his corn popping!

As soon as Lieutenant Cornlumbo mentioned fat determination , Shallot Holmes became very interested. “Now we’re talking,” said Holmes. And he went on to question just how Cornlumbo had gone about determining his fat content. Cornlumbo, who loved nothing more than writing a list, had made notes detailing his process and methods of determination. He pulled out his notes and started to explain how he had got his results. Shallot Holmes noticed with his keen eye for detail that several results were crossed out in his notes. “Do you mind if I take a look at your method and results, Lieutenant Cornlumbo?” asked Holmes. Cornlumbo looked nervous as he handed his notes over for inspection.

Shallot Holmes took a seat and started his investigation. The detectives sat anxiously watching as Shallot tutted and scratched his head while reading. Having finished reading the notes, Shallot Holmes rose to his feet and declared, “Detection is, or ought to be, an exact science and should be treated in the same cold and unemotional manner.” He pointed out that although Cornlumbo had done repeat experiments, his results varied considerably, and only the lowest numbers had been included in his conclusion. High Standard deviations are a clear indication that the method used was not consistent or well-considered. If the samples were not homogenous, this may have caused the standard deviations, but I fear this was not the only error made. Lieutenant Cornlumbo looked a little flustered and embarrassed and proudly stated, “My wife says I’m the second smartest guy she knows.”

“She also claims there are 80 guys tied for first place,” quietly muttered Miss Mapple to Nancy Beef, who almost choked on her coffee.

Shallot Holmes continued in his assessment, noticing the amount of siphoning performed by Cornlumbo. “It looks as if you did not reach the point of practical conclusion, Lieutenant”. Holmes explained that this depended on the solubility of the substance being extracted. He saw Cornlumbo was looking quite disillusioned and made him feel better by explaining it often took a lot of experience to assess if this point had been reached. “Another way to determine the end of the extraction is to verify the method by using reference material with known fat content”.

“I also see that you did not perform a hydrolysis step at the beginning, so you only determined the free fat content of your sample.” Holmes described the difference between free and total fat detection. Total fat determination not only detects the free fat but also the fat enclosed by other components of the sample matrix. In baked goods, fat is often mechanically enclosed by carbohydrates to form glycolipids and protein substances to form lipoproteins. In dairy products, surface tension forces cause colloidal components to surround the fat droplets. In yeast – and our good friend Eggcule Poirot – certain fat components are bound chemically or by absorption, forming phosphatide-protein complexes. To release all these fats, the bonds that hold lipid and non-lipid components together must be broken down before the extraction process. During hydrolysis, proteins are hydrolyzed, and plant cell walls are broken down, releasing physically enclosed fats. “It is for these reasons that Total Fat Determination is the required method for compliance with several industry standards and very important for our clients,” said Holmes.

Holmes then asked the detectives if they knew the 3 primary methods used for fat determination. Miss Mapple was the first to answer as she was quite the expert on fat extraction as she had previously given five tips to optimize fat extraction. “Soxhlet extraction,” she said without hesitation and went on to describe how this method was used to determine fat content in dry solid samples. Holmes was very impressed. Eggcule Poirot was next to speak. He said that continuous extraction could also be used and offered high efficiency and accelerated analyte-solvent exchange due to higher temperatures than Soxhlet. “Eggcelent deduction,” said Holmes cheekily. The last method was described by Nancy Beef, who was reminded of it when Eggcule mentioned temperature. “The last method is hot extraction,” she said. This method she described as the Randall method, which involved placing the sample in boiling solvent. Holmes was again impressed by the knowledge of his fellow detectives. He said that Continuous and Soxhlet extraction methods are the easiest to set up and generally offer the most reproducible results. There was even now an instrument that allowed for all three methods offering a huge degree of versatility.

Holmes concluded by listing all of the areas that could negatively impact fat determination. The drying of the extract may not be sufficient; the fat could get oxidized due to excessively high temperatures before the drying step. There may be impurities in the solvent, which itself could be evaporated too fast. The sample may be too inhomogeneous and require a powerful blender to ensure homogeneity. For Soxhlet extraction, the number of cycles is essential, whereas the extraction time is the most critical factor for hot and continuous extraction. Holmes finished his explanation by saying that an accurate analysis of the Lieutenant’s post-Christmas fat content would involve doing a more thorough and accurate fat analysis using the advice and knowledge they had determined in the day’s discussion. The detectives, however, did not need to wait for the results of a subsequent test to determine that a diet of over 8000 calories a day was not ideal. They all made New Year’s resolutions to do more exercise and be aware of their food intake. Apart from Lieutenant Cornlumbo, who was looking out the window and thinking about chocolate.