Salivary Cystatin SN Binds to Phytic Acid In Vitro and Is a Predictor of Nonheme Iron Bioavailability with Phytic Acid Supplementation in a Proof of Concept Pilot Study

Abstract

Background Acute phytic acid intake has been found to decrease iron bioavailability; however, repeated phytic acid consumption leads to iron absorption adaptation. Salivary proline-rich proteins (PRPs) have been shown to inhibit iron chelation to tannins and may mediate similar iron absorption adaptation with phytic acid intake. Objectives The objectives of this study were to determine whether salivary proteins bind to phytic acid in vitro, and to explore a proof of concept in a pilot study that examined the impact of 4-wk, daily phytic acid supplementation on individuals’ iron status, bioavailability, and salivary PRP concentrations. Methods High-performance liquid chromatography (HPLC) and matrix-assisted laser desorption/ionization–time of flight were used to characterize in vitro salivary protein–phytic acid interactions. Nonanemic women ( n = 7) consumed 350 mg phytic acid supplements 3 times daily for 4 wk, and meal challenges were employed to determine iron bioavailability, iron status, and salivary protein concentrations before and after supplementation periods. Enzyme-linked immunosorbent assay (ELISA) analysis of purified protein fractions and participant saliva identified proteins bound to phytic acid. Results In vitro salivary protein–phytic acid interaction identified cystatin SN, a non–proline rich salivary protein, as the specific bound protein to phytic acid. Iron bioavailability ( P = 0.32), hemoglobin ( P = 0.72), and serum ferritin ( P = 0.08) concentrations were not reduced from week 0 to week 4 after phytic acid supplementation. Basic PRPs and cystatin SN concentrations were positively correlated with iron bioavailability at week 4. Conclusions Overall, results suggest that phytic acid binds to the non-PRP cystatin SN and that salivary protein production may improve iron bioavailability with phytic acid consumption.

Food Pattern Modeling as an Alternative Assessment Method to Multiday Dietary Recalls for Iron-Related Nutrients: A Proof-of-Concept Study

“…In this study, food pattern modeling was used to predict hematological indices for four variables: iron, ascorbic acid, phytate, and phytate–iron ratios. These variables were chosen due to their known impact on the hematological indices of iron status and to their common focus in community nutrition research and programming [ 3 , 4 , 5 , 6 , 7 , 8 , 9 , 10 , 11 , 12 , 13 , 14 , 17 , 18 , 19 , 21 , 22 , 23 , 24 , 27 ].…”

“…This suggests that, while food pattern modeling may be non-inferior to 24 h dietary recall methods for long-term markers of iron status, it may be inadequate to predict acute markers of iron absorption. However, it has been suggested that dietary iron intake may be poorly predicted by single-meal iron bioavailability studies compared to long-term markers or iron status [ 3 , 12 , 13 , 33 ], questioning whether the improved diagnostic capabilities of 24 h dietary recalls may be needed in community-based dietary assessment. In addition, the mean difference in diagnostic accuracy for all nutrient variables for any hematological outcome between assessment methods was less than twenty percent, often trending in favor of food pattern modeling methods for assessment.…”

“…To evaluate dietary recall methods against hematological outcomes, each participant completed an iron-rich meal challenge to assess for iron status and bioavailability. Meal challenges followed a format previously described [ 12 , 13 ]. Prior to the meal, two separate blood samples were collected by venipuncture in a 5-mL serum separator and 3-mL ethylenediaminetetraacetic acid (EDTA) evacuated tubes to measure serum iron (by spectrophotometry), ferritin (by immunoassay, sensitivity 0.1 ng/mL), and complete blood count, including hemoglobin (g/dL) [ 12 , 13 ].…”

“…All meals consisted of a 95-g bagel with 12 g sugar-free strawberry jam (half sprinkled with 18 mg iron as ferrous sulfate), a 90-g banana, and 355 mL of water. Blood samples were drawn to measure acute iron absorption at 210 min post-meal in a 2-timepoint draw to estimate the percentage of maximum iron recovery (% max iron absorption) as described previously [ 12 , 13 , 14 ]. After study activities were completed, blood and serum were sent to Quest Diagnostics for analysis within 24 h. Serum iron data were used to calculate acute iron absorption for iron bioavailability analysis.…”

“…After study activities were completed, blood and serum were sent to Quest Diagnostics for analysis within 24 h. Serum iron data were used to calculate acute iron absorption for iron bioavailability analysis. Calculations were completed as described previously [ 12 , 13 , 14 ].…”