Introduction

1. Identification of ACE-inhibitory precursor peptides from fermented camel milk (Camelusdromedarius) produced by Lactobacillus helveticus or Lactobacillus acidophilus using HPLC-MS Alhaj O. A**, Brückner, H* and Al-Khalifa, A. S; Department of Food Science & Nutrition, College of Food and Agricultural Sciences, King Saud University, Riyadh, Kingdom of Saudi Arabia, P.O. Box 2460, Riyadh 11451. *Visiting Professor; **oalhaj@ksu.edu.sa Introduction Aims Conclusion Camel milk has an important role in human nutrition in Arab countries andconsumed as fresh or soured milk (Al haj & AlKanhal, 2010). Saudi Arabia is the world’s second biggest camel milk producer with 89,000 tons per year (FAO, 2008). Hypertension is recognized as a serious risk factor for cardiovascular diseases, whereas one third of the Western population is estimated to have a risk factor for cardiovascular disease and stroke (López-Fandiño et al., 2006). The current ACE-inhibitory drugs include Captopril and Enalapril have some adverse side effects on patients including coughing, skin rashes, hypotension and increased potassium levels (Meisel et al., 2006). Pan et al., (2005) defined the angiotensin-I-converting enzyme (ACE; peptidyldipeptide hydrolase, EC 3.4.15.1) as “an exopeptidase that cleaves dipeptides from the C-terminal ends of various peptide substrates and regulates the activity of several endogenous bioactive peptides”. ACE-inhibitory peptides are present in the primary structure of various food proteins sources including milk protein (Li et al., 2004; Jang & Lee, 2005; Meisel et al. 2006). Daily ingestion of fermented bovine sour milk containing a starter culture of Lactobacillus (Lb.) helveticus produces ACE-inhibitory peptides. The information on camel milk health benefits are not. No research identified ACE-inhibitory peptides from dromedary camel milk proteins. Therefore, the aim of this research is to identify ACE-inhibitory peptides from dromedary camel milk β-casein produced by Lb. helveticus or Lb. acidophilus. HPLC-MS analysis of the fermented camel milk containing Lb. helveticus hydrolysed for 127h shown to have Tyr (Y), Arg (R) and Pro (P) at their ultimate C-terminal position, making them a possible candidate for ACE-inhibitory activity. These amino acids within the peptides were previously found to contribute substantially to ACE-inhibitory potency. Further investigation will be undertaken to study the direct ACE-inhibitory activity of fermented camel milk. • To investigate whether Lb.helveticus or Lb. acidophilus able to exhibit ACE-inhibitory activity from camel milk. • To identify ACE-inhibitory precursor peptides from fermented camel milk β-casein produced by Lb. helveticus or Lb. acidophilususing HPLC-MS. Results and Discussion Fermented camel milk was investigated using Lb. helveticus or Lb. acidophilus, but Lb. helveticus was shown to be superior in respect to production of ACE-inhibitory peptides. Seven ACE-inhibitory peptides were produced by Lb. helveticus from camel milk, while only two ACE-inhibitory peptides were produced by Lb. acidophilus probably due to its slower growth. All identified peptide sequences result from β-casein was found to be similar in their C-terminal position but different in their length. Corresponding amino acid sequences of peptides derived from fermented camel milk β-casein containing Lb. helveticus or Lb. acidophilus expected to have ACE-inhibitory activity References Alhaj O A and AlKanhal H A (2010). Compositional, Technological and Nutritional aspects of Dromedary Camel Milk - a Review. International Dairy Journal 20: 811-821. FAO. (2008) Camel milk. Retrievable from http://www.fao.org/ag/againfo/themes/en/dairy/camel.html Jang, A., & Lee, M. (2005). Purification and identification of angiotensin converting enzyme inhibitory peptides from beef hydrolysates. Meat Science69: 653-661. Li, G. H., Le, G. W., Shi, Y. H., & Shrestha, S. (2004). Angiotensin I-converting enzyme inhibitory peptides derived from food proteins and their physiological and pharmacological effects. Nutrition Research24: 469-486. López-Fandiño, R., Otte, J., & Van Camp, J. (2006). Review: Physiological, chemical and technological aspects of milk-protein-derived peptides with antihypertensive and ACE-inhibitory activity. International Dairy Journal16: 1277-1293. Meisel, H., Walsh, D. J., Murray, B. & FitzGerald, R. J. (2006). ACE inhibitory peptides. Chapter 13, pp 269-315. In Nutraceutical Proteins and Peptides in Health and Disease Vol. 4, (Edited by Mine Y and Shahidi F, Taylor & Francis Group Publisher). Pan, D., Luo, Y. & Tanokura, M. (2005). Antihypertensive peptides from skimmed milk hydrolysate digested by cell-free extract of Lactobacillus helveticus JCM1004. Food Chemistry91: 123–129. Methodology ACE-inhibitory Identified peptide Starter peptide (a) sequence (b) KVLPVP LSLSQFKVLPVPQLb. helveticus SLSQFKVLPVPQ SQFKVLPVPQ LHLPLP TDLENLHLPLPLLb. helveticus DLENLHLPLPL LENLHLPLPL AVPYPQR AVPYP KVLPVVPQQMVPYPQLb. helveticus PYP YQEPVLGPVR FQEPVPDPVRLb. acidophilus VLPFQEPVPDPVRG Whole camel milk was collected from dromedary camels (‘Majaheim’ breed) from a farm located in Al-Jawf city in the north region of Saudi Arabia. The pH and acidity of camel milk were 6.54 and 0.14%, respectively. Milk was sterilized using an Arnold steam sterilizer and then inoculated with Lb.helveticus (LMG11445) or Lb. acidophilus (LMG11430) strains and incubated anaerobically at 37C for 127h and 112h, respectively. Samples were filtered through 3k Da cutoff dialysis membrane and fat was removed from the permeate by extraction with hexane. Aliquots of 5ml were analyzed by HPLC-MS. Derived peptides masses were then identified and searched against Swissprot protein database using MASCOT. Acknowledgement This research is supported by The National Plan for Science & Technology (NPST) Program by King Saud University Project Number 11-AGR1603-02. (a) previously characterized as ACE-inhibitory peptides from bovine milk by Meisel et al. (2006); (b) sequences of peptides derived from fermented camel milk; underlined sequences in (b) correspond to those depicted in (a).